The design of more efficient photosensitizers is a matter of great importance in the field of cancer treatment by means of photodynamic therapy. One of the main processes involved in the activation of apoptosis in cancer cells is the oxidative stress on DNA once a photosensitizer is excited by light. As a consequence, it is a matter of great relevance to investigate in detail the binding modes of the chromophore with DNA, and the nature of the electronically excited states that participate in the induction of DNA damage, for example, charge-transfer states. In this work, we investigate the electronic structure of the anthraquinone photosensitizer intercalated into a double-stranded poly(dG-dC) decamer model of DNA. First, the different geometric configurations are analyzed by means of classical molecular dynamics simulations. Then, the excited states for the most relevant poses of anthraquinone inside the binding pocket are computed by an electrostatic-embedding quantum mechanics/molecular mechanics approach, where anthraquinone and one of the nearby guanine residues are described quantum mechanically to take into account intermolecular charge-transfer states. The excited states are characterized as monomer, exciton, excimer and charge-transfer states based on the analysis of the transition density matrix, and each of these contributions to the total density of states and absorption spectrum is discussed in terms of the stacking interactions. These results are relevant as they represent the footing for future studies on the reactivity of anthraquinone derivatives with DNA and give insights on possible geometrical configurations that potentially favor the oxidative stress of DNA.

Today massive amounts of sequenced metagenomic and -transcriptomic data from different ecological niches and environmental locations are available. Scientific progress depends critically on methods that allow extracting useful information from the various types of sequence data. Here, we will first discuss types of information contained in the various flavours of biological sequence data, and how this information can be interpreted to increase our scientific knowledge and understanding. We argue that a mechanistic understanding is required to consistently interpret experimental observations, and that this understanding is greatly facilitated by the generation and analysis of dynamic mathematical models. We conclude that, in order to construct mathematical models and to test mechanistic hypotheses, time-series data is of critical importance. We review diverse techniques to analyse time-series data and discuss various approaches by which time-series of biological sequence data was successfully used to derive and test mechanistic hypotheses. Analysing the bottlenecks of current strategies in the extraction of knowledge and understanding from data, we conclude that combined experimental and theoretical efforts should be implemented as early as possible during the planning phase of individual experiments and scientific research projects.

Directed evolution methods are becoming increasingly popular, as they are extremely powerful toward developing new biomolecules with altered/novel activities, e.g., proteins with new catalytic functions or substrate specificities, and nucleic acids that recognize an intended target. Especially useful are systems that have incorporated continuous evolution, where the protein to be evolved undergoes continuous mutagenesis to evolve a desired trait with little to no input from the researcher once the system is started. However, continuous evolution methods can be challenging to implement in the lab and daunting for researchers to invest time and resources. Our intent is to provide basic information and helpful suggestions that we have gained from our experience with bacterial phage-assisted continuous evolution (PACE). Specifically, we review factors to consider before adopting PACE for a given evolution scheme, different types of selection circuits that can be utilized with particular focus on the PACE-B1H selection system, what optimization of a PACE selection circuit may look like using directed evolution of ME47 as a case study, and additional techniques that may be incorporated into a PACE experiment. With this information, researchers will be better equipped to determine if PACE is a valid strategy to use to evolve their proteins and how to set up a valid selection circuit.

Epigenetic changes are a hallmark of short- and long-term transcriptional regulation, and hence instrumental in the control of cellular identity and plasticity. Epigenetic mechanisms leading to changes in chromatin structure, accessibility for recruitment of transcriptional complexes, and interaction of enhancers and promoters all contribute to acute and chronic adaptations of cells, tissues and organs to internal and external perturbations. Similarly, the peroxisome proliferator-activated receptor γ coactivator 1α (PGC-1α) is activated by stimuli that alter the cellular energetic demand, and subsequently controls complex transcriptional networks responsible for cellular plasticity. It thus is of no surprise that PGC-1α is under the control of epigenetic mechanisms, and constitutes a mediator of epigenetic changes in various tissues and contexts. In this review, we summarize the current knowledge of the link between epigenetics and PGC-1α in health and disease.

A platinum (II) complex stabilized by a pyridine and a N-heterocyclic carbene ligand featuring an anthracenyl moiety was prepared. The compound was fully characterized and its molecular structure was determined by single-crystal X-ray diffraction. The compound demonstrated high in vitro antiproliferative activities against cancer cell lines with IC50 ranging from 10 to 80 nM. The presence of the anthracenyl moiety on the NHC Pt complex was used as a luminescent tag to probe the metal interaction with the nucleobases of the DNA through a pyridine-nucleobase ligand exchange. Such interaction of the platinum complex with DNA was corroborated by optical tweezers techniques and liquid phase AFM microscopy. The results revealed a two-state interaction between the platinum complex and the DNA strands. This two-state behaviour was quantified from the different experiments due to contour length variations. At 24h incubation, the stretching curves revealed multiple structural breakages, and AFM imaging revealed a highly compact and dense structure of platinum complexes bridging the DNA strands.

Environmental DNA (eDNA) is an increasingly used non-invasive molecular tool for detecting species presence and monitoring populations. In this article, we review the current state of non-avian reptile eDNA work in aquatic systems, as well as present a field experiment on detecting the presence of painted turtle (Chrysemys picta) eDNA. Thus far, turtle and snake eDNA studies have been successful mostly in detecting the presence of these animals in field conditions. However, some instances of low detection rates and non-detection occur for these non-avian reptiles, especially for squamates. We explored this matter by sampling lentic ponds with different densities (0 kg/ha, 6 kg/ha, 9 kg/ha, and 13 kg/ha) of painted turtles over three months, attempting to detect differences in eDNA accumulation using a qPCR assay. Only one sample of the highest density pond readily amplified eDNA. Yet, estimates of eDNA concentration from pond eDNA were rank-order correlated with turtle density. We present a “shedding hypothesis”–the possibility that animals with hard, keratinized integument do not shed as much DNA as mucus-covered organisms–as a potential challenge for turtle eDNA studies. Despite challenges with eDNA inhibition and availability in water samples, we remain hopeful that eDNA can be used to detect freshwater turtles in the field. We provide key recommendations for biologists wishing to use eDNA methods for detecting non-avian reptiles.

Next-generation sequencing is now a mature technology, allowing partial animal genomes to be produced for many clades. Though many software exist for genome assembly and annotation, a simple pipeline that allows researchers to input raw sequencing reads in fastq format and allow the retrieval of a completely assembled and annotated mitochondrial genome is still missing. mitoMaker 1.0 is a pipeline developed in python that implements (i) recursive de novo assembly of mitochondrial genomes using a set of increasing k-mers; (ii) search for the best matching result to a target mitogenome and; (iii) performs iterative reference-based strategies to optimize the assembly. After (iv) checking for circularization and (v) positioning tRNA-Phe at the beginning, (vi) geneChecker.py module performs a complete annotation of the mitochondrial genome and provides a GenBank formatted file as output.

Cisplatin (CDDP), carboplatin (CP), and oxaliplatin (OXP) are three platinating agents clinically approved worldwide for use against a variety of cancers. They are canonically known as DNA damage inducers; however, that is only one of their mechanisms of cytotoxicity. CDDP mediates its effects through DNA damage-induced transcription inhibition and apoptotic signalling. In addition, CDDP targets the endoplasmic reticulum (ER) to induce ER-stress, the mitochondria via mitochondrial DNA damage leading to ROS production, and the plasma membrane and cytoskeletal components. CP acts in a similar fashion to CDDP by inducing DNA damage, mitochondrial damage, and ER stress. Additionally, CP is also able to upregulate micro-RNA activity, enhancing intrinsic apoptosis. OXP, on the other hand, at first induces damage to all the same targets as CDDP and CP, yet it is also capable of inducing immunogenic cell death via ER stress and can decrease ribosome biogenesis through its nucleolar effects. In this comprehensive review, we provide detailed mechanisms of action for the three platinating agents, going beyond their nuclear effects to include their cytoplasmic impact within cancer cells. In addition, we cover their current clinical use and limitations, including side effects and mechanisms of resistance.

The Asian hornet, Vespa velutina, is an invasive species that has not only expanded its range in Asia but also invaded European countries, incurring significant costs on the local honey and bee industry. This phylogeographic study aims to trace the evolutionary trajectory of V. velutina and its close relatives, aiming to identify features that characterize an invasive species. The last successful invasion of Vespa velutina into France occurred in 2002.40, and into South Korea in 2002.77, estimated by fitting a logistic equation to the number of observations over time. The instantaneous rate of increase is 1.3667 for V. velutina in France and 0.2812 in South Korea, consistent with the interpretation of little competition in France and strong competition from local hornet species in South Korea. The invasive potential of two sister lineages can be compared by their distribution area when proper statistical adjustments are made to account for differences in sample size. V. velutina has a greater invasive potential than its sister lineage. The ancestor of V. velutina split into two lineages, one found in Indonesia/Malaysia and the other colonizing the Asian continent. The second lineage split into a sedentary clade inhabiting Pakistan and India, and an invasive lineage colonizing much of Southeast Asia. This latter lineage gave rise to subspecies V. v. nigrithorax that invaded France, South Korea, and Japan. My software PGT, which generates geophylogenies and computes geographic areas for individual taxa, is useful for understanding biogeography in general and invasive species in particular. I discussed the conceptual formulation of an index of invasiveness for comparison between sister lineages.

Patients with heart failure (HF) with improved LVEF (HFimpEF) demonstrate better clinical outcomes when compared with individuals without restoration of cardiac function. Identification of predictors for HFimpEF may play a crucial role in individual management of HF with reduced EF (HFrEF). Cell-free nuclear (cf-nDNA) DNA are released from damaged cells and contribute to adverse cardiac remodeling, cardiac dysfunction and inflammation. The purpose of the study was to elucidate whether cf-nDNAs are associated with HFimpEF. It has been prescreened 1416 patients with HF using local database. Between October 2021 and August 2022 we included 452 patients chronic HFrEF after prescription of optimal guideline-based therapy and identified HFimpEF in 177 individuals from them according to the criteria of European Society of Cardiology. Measurements of circulating biomarkers were performed at baseline and in 6 months. Detection of cf-nDNA was executed with real time quantitative PCR (qPCR) using NADH dehydrogenase, ND2 and beta-2-microglobulin. We found that HFimpEF was associated with a significant decrease in the levels of cf-nDNA when compared with the patients from persistent HFrEF cohort. Presence of ischemia-induced cardiomyopathy (odds ration [OR] = 0.75; confidence interval [CI] = 0.62 – 0.88; p = 0.044), type 2 diabetes mellitus (OR = 0.77; CI = 0.71 – 0.82; p = 0.042) and digoxin administration (OR = 0.85; CI = 0.72-0.97; p = 0.042) were negative factors for HFimpEF, whereas NT-proBNP ≤1940 pmol/mL (OR = 1.42; 95% CI = 1.19-1.98, p = 0.001), relative decrease in NT-proBNP levels (>35% vs. ≤35%) from baseline (OR = 1.52; 95% CI = 1.38-0.69, p = 0.001) and cf-nDNA ≤7.5 μmol/L (OR = 1.56; 95% CI = 1.07-2.94, p = 0.001) were positive predictors for HFimpEF. Multivariate logistic regression adjusted to ischemia-induced cardiomyopathy, IV HF NYHA class, and digoxin use yielded that NT-proBNP ≤1940 pmol/mL (OR = 1.43; 95% CI = 1.21-1.88, p = 0.001) and cf-nDNA ≤7.5 μmol/L (OR = 1.64; 95% CI = 1.19-2.15, p = 0.001) independently predicted HFimpEF. In conclusions: we established that the levels of cf-nDNA≤7.5 μmol/L independently predicted HFimpEF and improved discriminative ability of as single measured NT-proBNP as well as relative decrease in NT-proBNP levels ≤35% from baseline.

Histone deacetylase inhibitors (HDACis) are being recognized as a potentially effective treatment approach for peripheral T-cell lymphoma (PTCL), which is a diverse group of aggressive lymphomas with a bleak prognosis. Recent evidence has shown that HDACi are effective in treating PTCL, especially in cases where the disease has relapsed or is resistant to other treatments, and there are few available options. Several clinical trials have demonstrated that HDACi, such as romidepsin and belinostat, can elicit long-lasting positive outcomes in individuals with PTCL, either when used alone or in conjunction with conventional chemotherapy. They exert their anti-tumor actions by regulating gene expression through inhibiting histone deacetylases, which results in the halting of cell division, programmed cell death, and the transformation of cancerous T-cell, as indicated by gene expression profile studies. Importantly, besides clinical trials, real-world evidence indicated that the utilization of HDACi presents a significant and beneficial treatment choice for PTCL. However, although HDACi showed potential effectiveness, they couldn’t cure most patients. Therefore, new combination with conventional drugs as well as new targeted agents are under investigation.

Parkinson’s disease (PD) is one of the most common neurodegenerative disorders globally, and leads to an excessive loss of dopaminergic neurons in the substantia nigra of the brain. Circulating cell-free DNA (ccf-DNA) are double-stranded DNA fragments of different sizes and origins that are released into the serum and cerebrospinal fluid (CSF) due to cell death (i.e., necrosis and apoptosis) or are actively released by viable cells via exocytosis and NETosis. Methods: Using droplet digital polymerase chain reaction (ddPCR) we comprehensively analyzed and distinguished ccfDNA and ccf mtDNA in the serum and CSF of PD and control patients. Results: The quantitative analysis of serum ccf-DNA in PD patients demonstrated significant increase of ccf mtDNA and ccfDNA compared to healthy control patients and significantly higher copy of ccf mtDNA when compared to ccfDNA. Next, the serum ccf mtDNA levels significantly increased in male PD patients compared to healthy male controls. Furthermore, CSF ccf mtDNA in PD patients increased significantly compared to ccfDNA, and ccf mtDNA decreased in PD patients compared to healthy controls. These decreases were not statistically significant, but were in agreement with previous data. In-terestingly, ccf mtDNA increased in healthy control patients in both serum and CSF as compared to ccfDNA. Limitations: the small sample size of serum and CSF were the main limitations of this study. Conclusion: To the best of our knowledge, this is the first comprehensive study on serum and CSF of PD patients using ddPCR and indicating the distribution of the copy number of ccf mtDNA, as well as ccfDNA. If validated, we suggest that ccf mtDNA has greater than ccfDNA potential to lead the development of novel treatments for PD patients.

Vaccine development against SARS-CoV-2 has been highly successful in slowing down the COVID-19 pandemic. A wide spectrum of approaches including vaccines based on whole viruses, protein subunits and peptides, viral vectors, and nucleic acids have been developed in parallel. For all types of COVID-19 vaccines, good safety and efficacy have been obtained in both preclinical animal studies and in clinical trials in humans. Moreover, emergency use authorization has been granted for all types of COVID-19 vaccines. Although high safety has been demonstrated, rare cases of severe adverse events have been detected after global mass vaccinations. Emerging SARS-CoV-2 variants possessing enhanced infectivity have affected vaccine protection efficacy requiring re-design and re-engineering of novel COVID-19 vaccine candidates. Furthermore, insight into preparedness against emerging SARS-CoV-2 variants and other future viral infections is given.

The APOE locus has garnered significant clinical interest because of its association with Alzheimer's disease (AD) and longevity. This genetic association appears across multiple genes in the APOE locus. Despite the apparent differences between AD and longevity, both conditions share a commonality of aging-related changes in mitochondrial function. This commonality is likely due to accumulative biological effects, partly exerted by the APOE locus. In this study, we investigated changes in mitochondrial structure/function-related markers using oxidative stress-induced human cellular models and postmortem brains (PMBs) from individuals with AD and normal controls. Our results reveal a range of expressional alterations, either up- or down-regulated, in these genes in response to oxidative stress. In contrast, we consistently observed an upregulation of multiple APOE locus genes in all cellular models and AD PMBs. Additionally, the effects of AD status on mitochondrial DNA copy number (mtDNA CN) varied depending on APOE genotype. Our findings imply a potential coregulation of APOE locus genes, possibly occurring within the same topologically associating domain (TAD) of the 3D chromosome conformation. The coordinated expression of APOE locus genes could impact mitochondrial function, contributing to the development of AD or longevity. Our study underscores the significant role of the APOE locus in modulating mitochondrial function and provides valuable insights into the underlying mechanisms of AD and aging, emphasizing the importance of this locus in clinical research.

Tungiasis is a neglected tropical disease caused by skin-penetrating female Tunga penetrans fleas. Although tungiasis causes severe health problems, its ecology is poorly understood and morphological descriptions of larvae are unavailable. To identify T. penetrans immature stages and sites where they develop, diagnostic PCRs are required. However, flea larvae feed on soil organic matter rich in PCR inhibitors. Here, three DNA preparation methods, a soil DNA kit removing inhibitors, a simple ammonium acetate precipitation approach (AmAcet) and a crude lysate of larvae (CL), were combined with amplification by the highly processive FIREPol® Taq or the inhibitor-resistant Phusion® polymerase. Independent of the polymerase used, frequency of successful amplification, Cq values and PCR efficacies for the low-cost CL and AmAcet methods were superior to the commercial kit for amplification of a 278 bp partial internal transcribed spacer-2 (ITS-2) and a 730 bp pan-Siphonaptera cytochrome oxidase I PCR. For the CL method combined with Phusion® polymerase, costs were approximately 20-fold lower than for methods based on the soil DNA kit, which is a considerable advantage in resource-poor settings. The ITS-2 PCR did not amplify Ctenocephalides felis genomic or Tunga trimammilata ITS-2 plasmid DNA allowing it to be used to specifically identify T. penetrans.

The majority of Candida species are known as non-pathogenic yeasts and rarely involved in human diseases. However, recently case reports of human infections caused by non-albicans Candida species have increased, mostly in immunocompromised hosts. Our study aimed to describe and caracterise as thoroughly as possible, a new species of the Candida genus, named here Candida massiliensis (PMML0037), isolated from a clinical sample of human sputum. We compared genetic data based on the sequences of four genetic regions: "Internal Transcribed Spacers" of rRNA, D1/D2 domains (28S large subunit rRNA) and part of the genes encoding Translation Elongation Factor 1-α and β-tubulin2, to morphological characters, from scanning electron microscopy (TM 4000 Plus, SU5000), physiological, including the results of oxidation and assimilation tests of different carbon sources by the Biolog system, and chemical mapping by Energy-Dispersive X-ray Spectroscopy. Lastly, the in vitro antifungal susceptibility profile was performed using the E-testTM exponential gradient method. The multilocus analysis supported the genetic position of Candida massiliensis (PMML0037) as a new species of the genus Candida, and the phenotypic analysis highlighted its unique morphological and chemical profile when compared to other Candida species included in the study.

Background: Transient receptor potential cation channel subfamily A member 1 (TRPA1) is expressed in trigeminal neurons and brain regions important in migraine pathogenesis and is activated by many migraine triggers. Epigenetic regulation of TRPA1 expression is important in pain transmission and neurogenic inflammation.Findings: TRPA1 channels change noxious stimuli into pain signals with the involvement of epigenetic regulation, including DNA methylation, histone modifications, and effects of micro RNAs (miRNAs) and long non-coding RNAs. TRPA1 may change epigenetic profile of many pain-related genes as it may modify enzymes establishing the epigenetic profile and expression of non-coding RNAs. TRPA1 may induce the release of calcitonin gene related peptide (CGRP), from trigeminal neurons and dural tissue. Therefore, epigenetic regulation of TRPA1 may play a role in efficacy and safety of anti-migraine therapies targeting TRP channels and CGRP. TRPA1 is also involved in neurogenic inflammation, important in migraine. The fundamental role of TRPA1 in inflammatory pain transmission may be epigenetically regulated. Conclusions: Epigenetic connections of TRPA1 may play a role in efficacy and safety of anti-migraine therapy targeting TRP channels or CGRP and they should be further explored for efficient and safe antimigraine treatment.

The article continues the author's publications about the matrix-tensor study of universal rules of stochastic (probabilistic) organization of long single-stranded DNA sequences in eukaryotic and prokaryotic genomes. The author reveals that corresponding matrices of probabilities of n-plets in n-textual representations of each genomic DNA are numerically interrelated each with other in such algebraic form, which has analogies with formalisms of the known tensor-matrix theory of digital antenna arrays. These arrays combine many separate antennas into a single coordinated ensemble with unique emergent properties, due to which antenna arrays are widely used in devices of medicine, astrophysics, avionics, etc. The noted analogies allow putting forward the author's hypothesis that the stochastic organization of genomic DNAs is connected with bio-antenna arrays. From the point of view of this hypothesis, many known facts about using principles of antenna arrays in inherited physiological phenomena are collected in a single grouping with genomic DNAs. This new topic about the biological meaning of profitable properties of antenna arrays includes problems of biological evolution, the origin of the genetic code, regenerative medicine, and the development of algebraic biology. These issues are discussed jointly with the author's results of quantum information analysis of stochastic features of genomic DNAs.

Heteromorphic sex chromosomes are rarely found in plants. They were observed only in 47 species from phylogenetically distant families, suggesting that the evolution of sex chromosomes was independent in these species. It was shown that DNA repeat sequences are one of the major factors driving sex chromosomes evolution, and an accumulation or elimination of the repetitive DNA elements are closely linked with the formation of differences in the sex chromosomes. The goal of this study was to characterize the transposon composition in male and female plants of Cannabis sativa L., Humulus lupulus L. and Humulus japonicus Siebold & Zucc. For the first time, the male and female genomes of H. japonicus as well as male genomes of H. lupulus and C. sativa have been sequenced (there were no open data about them). The analysis of genome-wide sequencing data with using Repeatexplorer2 and author’s scripts was carried out. It was shown that accumulation of Ty3-gypsy may be associated with speciation in Cannabaceae family which is the opposite of the theory of speciation throw whole-genome duplication. Moreover, the sex-specific DNA repeat clusters in C. sativa and H. japonicus were found. The analysis also revealed that the concentration of Tekay, Retand and Ikeros repeats in the Y chromosome of C. sativa is lower than in the X chromosome and the Angela concentration is higher in the Y chromosome.

Approximately 240 million people are chronically infected with hepatitis B virus (HBV), despite four decades of an effective HBV vaccine. During chronic infection, HBV forms two distinct templates responsible for viral gene transcription: (1) episomal covalently closed circular (ccc)DNA and (2) host-genome integrated viral templates. Multiple ubiquitous and liver-specific transcription factors are recruited onto these templates and modulate viral gene transcription. This review details the latest developments in antivirals that inhibit HBV gene transcription, and their impact on the stability of viral transcripts. Notably, nuclear receptor agonists exhibit potent inhibition of viral gene transcription from cccDNA, small molecule inhibitors repress HBV X protein-mediated transcription from cccDNA and small interfering RNAs and single-stranded oligonucleotides result in transcript degradation from both cccDNA and integrant templates. These antivirals mediate their effects by reducing viral transcripts abundance, eventually leading to loss of surface antigen expression, and can potentially be added to the arsenal of drugs with demonstrable anti-HBV activity. Thus, these candidates deserve special attention for future repurposing or further development as anti-HBV therapeutics.

The article presents the author's results of studying hidden rules of structural organizations of long DNA sequences in eukaryotic and prokaryotic genomes. The results concern some rules of percentages (or probabilities) of n-plets in genomes. To reveal such rules, the author considers genomic DNA nucleotide sequences as multilayers sequences of n-plets and studies the percentage contents of n-plets in different layers. Unexpected rules of invariance of total sums of percentages in certain tetra-groupings of n-plets in different layers of genomic DNA sequences are revealed. These discovered rules are candidates for the role of universal genomic rules. A tensor family of matrix representations of interrelated DNA-alphabets of 4 nucleotides, 16 doublets, 64 triplets, and 256 tetraplets is used in the study. This matrix approach allows revealing algebraic properties of the mentioned genetic rules of probabilities, which are useful for developing algebraic and quantum biology. Some analogies of the discovered genetic phenomena with phenomena of Gestalt psychology are noted and discussed. The author connects the received results about the genomic percentages rules with a supposition of P. Jordan, who is one of the creators of quantum mechanics and quantum biology, that life's missing laws are the rules of chance and probability of the quantum world. Additional attention is paid to the algebraic features of the system of structured DNA alphabets and their relationship with the methods of algebraic holography, known in the technique of processing discrete signals. The concept of algebraic-holographic genetics is being developed for the understanding of inherited holographic properties of organisms.

Ostreococcus tauri is a simple unicellular green alga representing an ecologically important group of phytoplankton in oceans worldwide. Modern molecular techniques must be developed in order to understand the mechanisms that permit adaptation of microalgae to their environment. We present for the first time in O. tauri a detailed characterization of individual genomic integration events of foreign DNA of plasmid origin after PEG-mediated transformation. Vector integration appears to be random, occurring mainly at a single locus, and thus confirming the utility of this technique for insertional mutagenesis. While the mechanism of double-stranded DNA repair in the O. tauri model remains to be elucidated, we clearly demonstrate by genome resequencing that the integration of the vector leads to frequent structural variations (deletions/insertions and duplications) and some chromosomal rearrangements in the genome at the insertion loci, and often within the vector sequence itself. From these observations, we speculate that a non-homologous end joining-like mechanism is required during random insertion events, as described in plants and other freshwater algal models. PEG-mediated transformation is therefore a promising molecular biology tool, not only for functional genomic studies, but also for biotechnological research in ecologically important marine algae.

The author's method of oligomer sums for analysis of oligomer compositions of eukaryotic and prokaryotic genomes is described. The use of this method revealed the existence of general rules for cooperative oligomeric organization of a wide list of genomes. These rules are called hyperbolic because they are associated with hyperbolic sequences including the harmonic progression 1, 1/2, 1/3, .., 1/n. These rules are demonstrated by examples of quantitative analysis of many genomes from the human genome to the genomes of archaea and bacteria. The hyperbolic (harmonic) rules, speaking about the existence of algebraic invariants in full genomic sequences, are considered as candidates for the role of universal rules for the cooperative organization of genomes. The described phenomenological results were obtained as consequences of the previously published author's quantum-information model of long DNA sequences. The oligomer sums method was also applied to the analysis of long genes and viruses including the COVID-19 virus; this revealed, in characteristics of many of them, the phenomenon of such rhythmically repeating deviations from model hyperbolic sequences, which are associated with DNA triplets. In addition, an application of the oligomer sums method are shown to the analysis of the following long sequences: 1) amino acid sequences in long proteins like the protein Titin; 2) phonetic sequences of long Russan literary texts (for checking of thoughts of many authors that phonetic organization of human languages is deeply connected with the genetic language). The topics of the algebraic harmony in living bodies and of the quantum-information approach in biology are discussed.

The advent of RNA-sequencing (RNA-Seq) technologies has markedly improved our knowledge and expanded the compendium of small non-coding RNAs, most of which derive from the processing of longer RNA precursors. In this review article, we will discuss about the biogenesis and function of small non-coding RNAs derived from eukaryotic ribosomal RNA (rRNA), called rRNA fragments (rRFs), and their potential role(s) as regulator of gene expression. This relatively new class of ncRNAs remained poorly investigated and underappreciated until recently, due mainly to the a priori exclusion of rRNA sequences—because of their overabundance—from RNA-Seq datasets. The situation surrounding rRFs resembles that of microRNAs (miRNAs), which used to be readily discarded from further analyses, for more than five decades, because we could not believe that RNA of such a short length could bear biological significance. As if we had not yet learned our lesson not to restrain our investigative, scientific mind from challenging widely accepted beliefs or dogmas, and from looking for the hidden treasures in the most unexpected places.

Schistosoma infection in snails can be monitored by microscopy or indirectly by sentinel mice. As both these approaches sometimes miss infections, more sensitive tests are needed, particularly in low-level transmission settings. In this study, the loop-mediated isothermal amplification (LAMP) technique, designed to detect a specific 28S ribosomal S. japonicum gene with high sensitivity, was compared to microscopy using snail samples from 51 areas endemic for schistosomiasis in five Chinese provinces. The results were compared with those by polymerase chain reaction (PCR) adding DNA sequencing as a reference when needed. The testing of pooled snail samples showed that a dilution factor of 1/50, i.e., one infected snail plus 49 non-infected ones, would still result in a positive reaction after the recommended number of amplification cycles. Testing a total of 232 pooled samples, emanating from 4,006 snail specimens, with the LAMP assay showed a 6.5% rate of infection, while traditional microscopy found only 0.04% positive samples in the same materials. Parallel PCR analysis confirmed the diagnostic accuracy of the LAMP assay, with DNA sequencing even giving LAMP a slight lead. Microscopy and the LAMP test were carried out at local schistosomiasis-control stations demonstrating that the potential of the latter assay to serve as a point-of-care (POC) test with results available within 60–90 minutes, while the more complicated PCR test had to be carried out at the National Institute of Parasitic Diseases (NIPD) in Shanghai, China. In conclusion, LAMP was found to be clearly superior to microscopy and as good as, or better, than PCR. Application of LAMP testing would be useful for surveillance and risk prediction as it requires less time than other techniques and can be used under field conditions, which improves and accelerates schistosomiasis control.

The invasion of the golden mussel has resulted in considerable environmental and socioeconomic alterations, which present a considerable threat to the native biodiversity and sustainability of the region. It is of the utmost importance to gain an understanding of the distribution and biological characteristics of this Asian mussel, as well as its interaction with human activities, in order to develop effective strategies for mitigating and preventing its further spread. This study examines the dispersal route and incidence of golden mussels, tracing their movement from initial populations in Argentina to their arrival in the São Francisco River Basin (SFR). The presence of the mussel was confirmed through an integrative assessment, which included shell taxonomic analyses and mitochondrial DNA signatures. This identified populations located 7.5 km from the river's mouth, in proximity to the Atlantic Ocean, in areas such as shrimp farms, artisanal ports, and marinas. The analysis of mitochondrial DNA revealed the presence of South American-specific and shared ancestral haplotypes in the SFR, Grande River, and Argentina. These findings indicate that intracontinental colonisation towards the northeast of South America originated from Asian populations that entered South America via Argentina. The absence of Asian-specific signatures in the RSF, combined with its geomorphological structure, which is unsuitable for large ports or transoceanic vessels, supports the hypothesis of intercontinental dispersal of Limnoperna fortunei.

Water samples for bacterial microbiome studies undergo a biomass concentration, DNA extraction and taxonomic identification steps. Through benchmarking, we studied the applicability of Skimmed milk flocculation (SMF) for bacterial enrichment, an adapted in-house DNA extraction protocol, and six 16S rRNA databases (16S-DBs). Surface water samples from two rivers were treated with SMF and Vacuum filtration (VF) and subjected to amplicon or shotgun metagenomics. A mock community standard underwent five DNA extraction protocols, taxonomical identification with six different 16S-DBs and evaluated by the Measurement Integrity Quotient (MIQ) score. In SMF samples, the skimmed milk was metabolized by members of lactic acid bacteria or genera such as Polaromonas, Macrococcus, and Agitococcus, resulting in increased relative abundance (p < 0.5) up to 5.0 log fold change compared to VF, rendering SMF inapplicable for bacterial microbiome studies. The best performing DNA extraction protocols were FastSpin Soil, the in-house method and EurX. All 16S-DBs yielded comparable MIQ scores within each DNA extraction kit, ranging from 61-66 (ZymoBIOMICs) up to 80-82 (FastSpin). DNA extraction kits exert more bias to the composition than the 16S-DBs. This benchmarking study presented valuable insights to inform future water metagenomic study designs.

Medulloblastoma (MB) is one of the most common malignant pediatric brain tumors. Current therapy results in poor prognosis for high-risk SHH/p53-mutated MB, emphasizing the importance of more effective therapeutic strategies. Here, we investigated potential radiosensitizing effects of checkpoint kinase inhibitors (Chk-i) prexasertib (Chk1/2) and SAR-020106 (Chk1) in human SHH/p53-mutated MB in vitro and in vivo. UW228 and DAOY cells were treated with Chk-i and irradiation (RT). Metabolic activity, proliferation, and apoptosis was determined at d3, long-term clonogenicity at d14. DNA damage was assessed after 1, 24, and 72 h. Patient-derived SHH/p53-mutated, luciferase-transfected MB cells were implanted orthotopically into NSG mice (d0). Fractionated therapy (daily, d7-11) was applied. Body weight (BW) was documented daily, tumor growth weekly, and proliferation at d42. In vitro, Chk-i exhibited dose-dependent reduction of metabolic activity, proliferation, and clonogenicity, and enhancement of apoptosis. Combination with RT showed additive antitumor effects and enhanced residual DNA damage. In vivo, tumor growth was delayed by Chk-i alone. Low-dose prexasertib enhanced RT-induced tumor growth inhibition (37-fold) and reduced proliferating cells, high-dose prexasertib and SAR-020106 showed opposite effects (n=3, n.s.). BW assessments revealed that the treatment was well tolerated. Our data indicate a benefit of Chk-i in combination with RT in SHH/p53-mutated MB. However, high-dose Chk-i may compromise the RT effect, possibly by its anti-proliferative activity. Furthermore, we demonstrate for the first time the antitumor activity of the Chk1-specific inhibitor SAR-020106 in the murine brain.

Sepsis is a life-threatening syndrome triggered by infection and accompanied by high mortality, with antimicrobial resistances (AMRs) further escalating clinical challenges. Rapid and reliable detection of causative pathogens and AMRs are key factors for fast and appropriate treatment, in order to improve outcome in septic patients. However, current sepsis diagnostics based on blood culture is limited by low sensitivity and specificity while current molecular approaches fail to enter clinical routine. Therefore, we developed a Suppression PCR-based selective enrichment sequencing approach (SUPSETS), providing a molecular method combining multiplex suppression PCR with Nanopore sequencing to identify most common sepsis-causative pathogens and AMRs using plasma cell-free DNA. Applying only 1 mL of plasma, we target eight pathogens across three kingdoms and ten AMRs in a proof-of-concept study. SUPSETS was successfully tested on first ten clinical samples and revealed comparable results to clinical metagenomics while clearly outperforming blood culture. Several clinically relevant AMRs could be detected additionally. Furthermore, SUPSETS provided first pathogen and AMR-specific sequencing reads within minutes of starting sequencing, thereby potentially decreasing time to results to 11 - 13 hours and suggesting diagnostic potential in sepsis.

Beyond factors such as smoking and obesity, the risk of getting gastrointestinal cancer is passed down from parents to offspring, inherited by a gene mutation, predisposing them to develop cancer in their lifetime. Here we describe the clinical history of family members affected by gastrointestinal pathologies often leading to cancer. In the epidemiological study, the cohort of members of this family shared a high risk of developing gastrointestinal diseases. The subjects were monitored from May 2006 to December 2017 by collecting periodically clinical and endoscopic data, and determining both auto-modification of lymphocyte Poly(ADP-ribose)Polymerase as early signal of DNA damage, and erythrocyte membrane lipid composition (Fat Profile). Both biomarkers were demonstrated to be non invasive and easy tools, to monitor over time family members, first the oldest members (nine siblings), and thereafter their offspring, whose cell status evolved from physiological even to cancerous one. All subjects developed gastrointestinal pathologies of different kind and seriousness. Some diseases evolved to cancer, sometimes as a sudden and lethal event. The results of the two molecular approaches, agreed and even predicted the clinical and imaging paths. This cohort study supported the hypothesis that both noninvasive molecular analyses can predict altered cell states and support clinical and imaging tests.

WBAN is a blessing to mankind because it can monitor people's health and activities regardless of place or time. It brings feasibility in patients’ treatment. The main problems with WBAN, however, are related to the security and privacy of health data. To prevent data misuse, health information should be safeguarded. On the other hand, the medical professional needs to receive the patient's health information promptly. As security of health data impact human life greatly so secure an efficient method is required. Efficient cryptographic technique for data protection of wireless body area network is proposed. This research aims to provide data protection in WBAN to increase data security and efficient transmission. The suggested approach offers effective security with quick key generation, quick encryption, and secure data transmission across the network. The proposed methodology is considered to be secure and will consume less time. The proposed method further uses a less complex encryption algorithm. The less complex the approach is the higher performance it will provide in wireless network. The proposed methodology will prevent many attacks, which makes the proposed approach highly suitable for wireless body area network’s security.

Transitions from seed to seedling represent a critical developmental step in the life cycle of higher plants, dramatically affecting plant ontogenesis and stress tolerance. The release from dormancy to acquiring the germination ability is defined by a balance of phytohormones, with a substantial contribution of abscisic acid (ABA), which inhibits germination. We studied the embryonic axes of Pisum sativum L. before and after radicle protrusion. Our previous work compared RNA sequencing-based transcriptomics in the embryonic axes isolated before and after radicle protrusion. The current study aims to analyze ABA-dependent gene regulation during the transition of embryonic axes from germination to post-germination stage. First, we determined the levels of abscisates (ABA, phaseic acid, dihydrophaseic acid, and neo-phaseic acid) using the ultra-high-performance liquid chromatography-tandem mass spectrometry. Second, we made a detailed annotation of ABA-associated genes using RNA sequencing-based transcriptome profiling. Finally, we analyzed the DNA methylation pattern in the promoters of the PsABI3, PsABI4, and PsABI5 genes. We showed that changes in the abscisate profile are characterized by the accumulation of ABA catabolites, and the ABA-related gene profile is accompanied by the upregulation of genes controlling the seedling development and the downregulation of genes controlling the water deprivation. The expression of ABI3, ABI4 and ABI5, which encode the crucial transcription factors during late maturation, was downregulated by more than 20-fold, and their promoters exhibited a high level of methylation already at the late germination stage. Thus, although ABA remains important, other regulators seems to be involved in the transition from seed to seedling.

Bisphenol is a chemical substance widely used in plastic products and food containers. In this study, we observed a relationship between DNA methylation and atopic dermatitis (AD) in the peripheral blood mononuclear cells (PBMCs) of pregnant women exposed to bisphenol A (BPA) and its alternatives, bisphenol S (BPS) and bisphenol F (BPF). DNA methylation is an epigenetic mechanism that regulates gene expression, which can be altered by environmental factors, and affects the onset and progression of diseases. We found that genes belonging to the JAK-STAT and PI3K-AKT signaling pathways were hypomethylated in the blood of pregnant women exposed to bisphenols. These genes play important roles in skin barrier function and immune responses, and may influence AD. Therefore, we suggest that not only BPA but also BPS and BPF, which are used as alternatives, can have a negative impact on AD through epigenetic mechanisms.

Zinc is an important nutrient for plant growth and development. Its availability is influenced by zinc solubilizing microbes (ZSM). Effects of commonly promoted agronomic practices on the abundance of ZSM are so far not well understood. In this study, we assessed the effects of farmyard manure (FYM) application, either sole, or in combination with residue and/or inorganic fertilizer inputs on ZSM community structure using 11 treatments in a long-term (17 years) integrated soil fertility management experiment located in western Kenya, The study was conducted in 2019. Bacterial and fungal community composition (at 0-15 cm depth) was evaluated by amplicon sequencing on an Illumina Miseq platform following the manufacturer’s guidelines. Microbial diversity indices (Shannon, Simpson and Chao1) and statistical analyses were computed using R project packages. Putative ZSM (i.e., the ZSM generally considered to possess the zinc solubilizing capabilities) were clustered in two major clades based on either application or no application of FYM. Sole application of FYM significantly (P<0.05) increased the abundance of several ZSM under maize-tephrosia rotation. In addition, systems with combined application of FYM with other inputs generally showed significantly increasing trends for some ZSM under maize-tephrosia rotation. Moreover, combined application of FYM and P rather than P only significantly increased abundance of some ZSM under maize-monocropping systems. Furthermore, besides affecting ZSM abundance, soil chemical variables involving soil organic carbon (SOC), total N and Olsen P significantly increased with FYM application. This study indicated that management practices like application of FYM that increase SOC, and other soil chemical parameters, also/concomitantly increase ZSM abundance; implying enhanced capacities for microbial-linked zinc availability.

Several tumor types have been efficiently treated with PARPis, which are now approved for the treatment of ovarian, breast, prostate, and pancreatic cancer. The BRCA1/2 genes and mutations in many additional genes involved in the HR pathway may be responsible for the HRD phenomenon. In the present study, an NGS assay was used to analyze 513 genes associated with targeted and immuno-oncology therapies in 406 samples. In addition, the %gLOH values of 24 samples were also calculated using the Affymetrix technology to compare the results obtained by the two methodologies. HR variations occurred in 20.93% of the malignancies, while BRCA1/2 gene alterations in 5.17%. %LOH was highly correlated with alterations in the BRCA1/2 genes since 76.19% (16/21) of the BRCA1/2 positive tumors had a high %LOH value (p=0.007). Moreover, the LOH status was highly correlated with the TP53 and KRAS statuses, but there was no association with the TMB value. Lin's Concordance Correlation Coefficient for the 24 evaluable samples simultaneously examined by both assays was 0.87, indicating a nearly perfect agreement. In conclusion, the addition of gLOH analysis could assist the detection of additional patients eligible for PARPis

Autosomal dominant isolated foveal hypoplasia (FVH1) is a rare condition of foveal hypoplasia (FH) that lacks any other ocular manifestations. FVH1 is associated with hypomorphic mutations in the PAX6 gene. We report our findings in 17 patients with PAX6 mutations associated with FVH1 or FH with aniridia and corneal opacities. Patients with three mutations, p.V78E, p.V83F and p.R128H, in the C-terminal subdomain of the paired domain (CTS) consistently had severe FH. Luciferase assays indicated that the transcriptional activities of these mutations were significantly reduced comparable to that with truncation mutation of p.G65Rfs*5. Patients with p.P20S in the N-terminal subdomain of the paired domain, and a patient with p.N365K in the proline-serine-threonine-rich domain (PSTD) had mild FH. A patient with p.Q255L in the homeodomain had severe FH. The P20S and Q255L mutants did not affect the transcriptional activity. Mutant N365K had a retained DNA binding activity but a reduced transcriptional activity due to a low PSTD transactivation. These findings demonstrated that mutations associated with FVH1 underlie a functional divergence between DNA binding ability and transcriptional activity. Broadly distributed mutations in the PAX6 gene, not limited to the CST region, were responsible for FVH1.

Cyclic vomiting syndrome (CVS) is a frequently disabling condition defined by severe, discrete, stereotypical episodes of nausea and vomiting. In our previous report [1], a candidate gene list was evaluated in a group of 80 unrelated participants with CVS with extensive genetic sequencing (whole exome/genome). Twenty-two good candidate genes for CVS association were identified, from which a cellular model of aberrant cation transport and energy metabolism was proposed. Herein, we present additional clinical information on those 80 participants, including eight case reports to highlight general principles and clinical practices. Our participants demonstrate substantial phenotypic and genotypic heterogeneity in CVS, including a propensity for specific co-morbidities in the participants and relatives, evolution towards episodes in which vomiting is less apparent, labeling with “non-organic” diagnoses, multiple genes, polygenic inheritance, and association with variants not designated as Pathogenic. Our case reports illustrate how genetic information can guide clinical management and argue to the high clinical utility of appropriate genetic testing and analysis in CVS. Expression profiling of our 22 candidate genes suggests an anatomical model in a unit defined by vagal afferents and adjacent cells. Our models are consistent with multiple current hypotheses of CVS and the efficacy of commonly-recommended treatments.

Rickettsia is an important pathogenic entity among tick-borne diseases (TBD), which are considered serious emerging public health problems globally. In India, though the widespread distribution of ticks and TBD has been documented, its real burden remains underreported. In a preliminary attempt, rickettsial surveillance was carried out in ticks collected from Sirumalai, Eastern Ghats in Tamil Nadu by using pathogen genome-based phylogenetic inferences generated through multi-locus sequence typing (MLST), targeting the genes 16s rRNA, OmpA, OmpB, and gltA by nested PCR. The laboratory evidence confirms the circulation of Rickettsia in Haemaphysalis intermedia species collected from this area. Analysis of the four gene sequences detected demonstrates their closest identity to the spotted fever group (SFG) available in the GenBank database. Further, multiple sequence alignment with other sequences derived from the GenBank database showed close relatedness to Rickettsia conorii subsp. raoultii (16s rDNA-99.32%, ompA-93.38%, ompB-97.39%, and gltA-98.57%) and Rickettsia felis (ompA-100%, ompB-100% and gltA-99.41%). With this genomic evidence, the circulation of rickettsial pathogens in the pools of H. intermedia ticks infesting livestock in the Sirumalai foothill area has been demonstrated and to complement the microscopic identification of the tick species, DNA barcodes were generated for H. intermedia using the mitochondrial cytochrome c oxidase subunit I gene (COI). Nevertheless, R. raoultii and R. felis were found to be the aetiological agents of tick-borne lymphadenopathy and flea-borne spotted fever in human cases, respectively. Further study on the determination of their diversity, distribution, clinical relevance, and potential risk to the local community in these areas is highly warranted.

Metal complexes feature a wide range of available geometries, diversified lability, controllable hydrolytic stability, and easily available rich redox activity. These characteristics combined with the specific properties of the coordinated organic molecules result in many different mechanisms of biological action, making each of the myriads of the classes of metal coordination compounds unique. This focused review presents combined and systematized results of the studies of a group of copper(I) (pseudo)halide complexes with aromatic diimines and tris(aminomethyl)phosphines of a general formula [CuX(NN)PR₃], where X = I^- or NCS^-, NN = 2,2’-bipyridyl, 1,10-phenanthroline, 2,9-dimethyl-1,10-phenanthroline or 2,2’-biquinoline, and PR₃ = Air-stable tris(aminomethyl)phosphines. The structural and electronic properties of the phosphine ligands and luminescent complexes are discussed. The complexes with 2,9-dimethyl-1,10-phenanthroline, apart from being air- and water-stable, exhibit a very high in vitro antimicriobial activity against the Staphylococcus aureus and Candida albicans. Moreover, some of the complexes also show a strong in vitro antitumor activity against human ovarian carcinoma cell lines: MDAH 2774 and SCOV 3, CT26 (mouse colon carcinoma), and A549 (human lung adenocarcinoma) cell lines. The tested complexes are moderately able to induce DNA lesions through free radical processes, however the trends do not reflect observed differences in biological activity.

AbstractPolyamines (PAs) including putrescine (PUT), spermidine (SPD) and spermine (SPM) are small, versatile molecules with two or more positively charged amino groups. Despite their importance for almost all forms of life, their specific roles in molecular and cellular biology remain partly unknown. The molecular structures of PAs suggest two presumable biological functions: (i) as potential buffer systems and (ii) as interactants with poly-negatively charged molecules like nucleic acids.The present report focuses on the question, whether the molecular structures of PAs are essential for such functions, or whether other simple molecules like small peptides with closely spaced positively charged side chains might be suitable as well. Consequently, we created titration curves for PUT, SPD, and SPM, as well as for oligolysines like tri-, tetra-, and penta-lysine. None of the molecules provided substantial buffering capacity at physiological intracellular pH values. Apparently, the most important mechanism for intracellular pH homeostasis in neurons is not a buffer system but is provided by the actions of the sodium-hydrogen and the bicarbonate-chloride antiporters.In a similar approach we investigated the interaction with DNA by following the extinction at 260 nm when titrating DNA with the above molecules. Again, PUT and tri-lysine were not able to interact with herring sperm DNA, while SPD and SPM were. Obviously, the presence of several positively charged groups on its own is not sufficient for the interaction with nucleic acids. Instead, the precise spacing of these groups is necessary for biological activity.

Attempts to find and quantify the supposed low entropy of organisms and its preservation are revised. Absolute entropy of the mixed components of non-living biomass (around -1.6 x 103 J K-1 L-1) is the reference to which other entropy decreases would be ascribed to life. Compartmentation of metabolites and departure from the equilibrium of metabolic reactions account for 1 and 40-50 J K-1 L-1, respectively, decreases of entropy and, though small, are distinctive features of living tissues. DNA and proteins do not supply significant decreases of thermodynamic entropy, but their low informational entropy is relevant for life and its evolution. No other living feature contributes significantly to the low entropy associated to life. The photosynthetic conversion of radiant energy to biomass energy accounts for the most of entropy (2.8 x 105 J K-1 carbon kg-1) produced by living beings. The comparative very low entropy produced in other processes (around 4.8 x102 J K-1 L-1 day-1 in human body) must be rapidly exported outside as heat to preserve the low entropy decreases due to compartmentation and non-equilibrium metabolism. Enzymes and genes are described whose control minimize the rate of production of entropy and could explain selective pressures in biological evolution and the rapid proliferation of cancer cells.

The aim of this study was to investigate the role of DNA methylation in the regulation of in vitro and in vivo cartilage formation. Based on the data of an RNA chip-assay performed on chondrifying BMP2-overexpressing C3H10T1/2 cells, the relative expression of Tet1 (tet methylcytosine dioxygenase 1), Dnmt3a (DNA methyltransferase 3) and Ogt (O-linked N-acetylglucosamine transferase) genes was examined with RT-qPCR in mouse cell-line based and primary micromass cultures. RNA probes for in situ hybridization were used on frozen sections of 15-day-old mouse embryos. DNA methylation was inhibited with 5-azacytidine during culturing. We found very strong but gradually decreasing expression of Tet1 throughout the entire course of in vitro cartilage differentiation along with strong signals in the cartilaginous embryonic skeleton. Dnmt3a and Ogt expressions did not show significant changes with RT-qPCR and gave weak in situ hybridization signals. Inhibition of DNA methylation applied during early stages of differentiation reduced cartilage-specific gene expression and cartilage formation. In contrast, it had stimulatory effect when added to differentiated chondrocytes. Our results indicate that the DNA demethylation-inducing Tet1 is a significant epigenetic factor of chondrogenesis, and inhibition of DNA methylation exerts distinct effects in different phases of in vitro cartilage formation.

The article shows materials to the question about algebraic features of the genetic code and about the dictatorial influence of the DNA and RNA molecules on the whole organism. Presented results testify in favor that the genetic code is an algebraic code related with a wide class of algebraic codes, which are a basis of noise-immune coding of information in communication technologies. Structural features of the genetic systems are associated with hypercomplex double (or hyperbolic) numbers and with bisymmetric doubly stochastic matrices. The received results confirm that represented matrix approaches are effective for modeling genetic phenomena and revealing the interconnections of structures of biological bodies at various levels of their organization. This allows one to think that living organisms are algebraically encoded entities where structures of genetic molecules have the dictatorial influence on inherited structures of the whole organism. New described algebraic approaches and results are discussed.

A large body of evidence reports about the positive effects of physical activity in pathophysiological conditions associated with aging. Physical exercise, alone or in combination with other medical therapies, unquestionably causes reduction of symptoms in chronic non-transmissible diseases often leading to significant amelioration or complete healing. The molecular basis of this exciting outcome, however, remain largely obscure. Epigenetics, exploring at the interface between environmental signals and the remodelling of chromatin structure, promises to shed light on this intriguing matter possibly contributing to the identification of novel therapeutic targets. In this review, we shall focalize on the role of sirtuins (Sirts) a class III histone deacetylases (HDACs) which function has been frequently associated, often with a controversial role, to the pathogenesis of aging-associated pathophysiological conditions including cancer, cardiovascular, muscular, neurodegenerative, bones and respiratory diseases. Numerous studies, in fact, demonstrate that Sirt-dependent pathways are activated upon physical and cognitive exercises linking mitochondrial function, DNA structure remodelling and gene expression regulation to designed medical therapies leading to tangible beneficial outcomes. However, in similar conditions, other studies assign to sirtuins a negative pathophysiological role. In spite of this controversial effect, it is doubtless that studying sirtuins in chronic diseases might lead to an unprecedented improvement of life quality in the elderly.

As with all new fields of discovery, work on the biological role of G-quadruplexes (GQ) has produced a number of results that at first glance are quite baffling, sometimes because they do not fit well together, but mostly because they are different from commonly held expectations. Like other classes of flipons, those that form G-quadruplexes have a repeat sequence motif that enables the fold. The canonical DNA motif (G3N1-7)3G3, where N is any nucleotide and G is guanine, a feature that is under active selection in avian and mammalian genomes. The involvement of G-flipons in genome maintenance traces back to the invertebrate Caenorhabditis elegans and to ancient DNA repair pathways. A role for GQ in transcription is supported by the observation that yeast Rap1 protein binds both B-DNA, in a sequence-specific manner, and GQ, in a structure-specific manner, through the same helix. Other sequence-specific transcription factors (TF) also engage both conformations to actuate cellular transactions. Noncoding RNAs can also modulate GQ formation in a sequence-specific manner and engage the same cellular machinery as localized by TF, linking the ancient RNA world with the modern protein world. The coevolution of noncoding RNAs and sequence-specific proteins is supported by studies of early embryonic development, where the transient formation of G-quadruplexes coordinates the epigenetic specification of cell fate.

Analysis of complex DNA mixtures is limited by the selected genetic markers characterized and assay technology. Small panels of short tandem repeats (STRs) are the current standards in most countries. Extension of STRs with hundreds of single nucleotide polymorphisms (SNPs) is now commercially available. The current set of extension SNPs focus on phenotypic and biogeographic ancestry informative SNPs. Larger panels with thousands of SNPs have been developed that can identify up to 10 contributors to forensic DNA samples. Herein, multiple sets of SNPs were selected with the focus of increasing the upper limit of detectable contributors. Larger SNP panel sets with lower minor allele frequency (MAF) alleles can detect large numbers of contributors to in silico mixtures of randomly selected profiles from the 1,000 Genomes project. These results provide a path to enable characterization of complex DNA mixtures that are currently impossible to analyse.

Psoralens have been used in the treatment of several skin diseases such as psoriasis and atopic eczema through PUVA (psoralen + UVA), cutaneous T-cell lymphoma through ECP (extracorporeal photochemotherapy), and recently, the treatment of solid tumors through X-PACT (x-ray psoralen activated cancer therapy). Ingesting psoralens and exposing the molecules to UVA light allows for the flat-planar psoralens to intercalate or slide in-between base pairs and then to form photo-adducts with the DNA backbone which can stop or slow DNA replication. The intrinsic ability of psoralens to bind with DNA and their therapeutic effectiveness contribute to the increased interest in the study of these molecules. Several studies measuring psoralen-DNA binding capabilities have used a variety of spectroscopic techniques and different experimental conditions. In this study, spectrofluorometry was used to measure the DNA-binding capabilities of three different psoralen derivatives (6E, AMT and 8-MOP) under identical experimental conditions. The binding constants were 0.325x106 M-1 for 8-MOP, 0.516x106 M-1 for AMT, and 7.30x106 M-1 for 6E. This direct comparison of the three different psoralens demonstrated the impact of their molecular structure on the DNA-binding capacity. 6E’s significantly greater binding constant makes it the stronger candidate for further studies looking into potential applications for cancer treatment.

In an era of increasing interest in the potential health benefits of medicinal foods, the need to assess their safety and potential toxicity remains a critical concern. While these natural remedies have garnered substantial attention for their therapeutic potential, a comprehensive understanding of their effects on living organisms is essential. We examined 316 herbal extracts to determine their potential nematocidal attributes in Caenorhabditis elegans. Approximately 16% of these extracts exhibited the capacity to induce diminished survival rates and larval arrest, establishing a correlation between larval arrest and overall worm viability. Certain extracts led to an unexpected increase in male nematodes, accompanied by a discernible reduction in DAPI-stained bivalent structures and perturbed meiotic advancement, thereby disrupting the conventional developmental processes. Notably, Onobrychis cornuta and Veratrum lobelianum extracts activated a DNA damage checkpoint response via the ATM/ATR and CHK-1 pathways; hindering germline development. Our LC‒MS analysis revealed jervine in V. lobelianum and nine antitumor compounds in O. cornuta. Interestingly, linoleic acid replicated phenotypes induced by O. cornuta exposure, including an increased level of pCHK-1 foci, apoptosis, and the MAPK pathway. We observed that mutants in the MAPK pathway suppressed the reduction in worm survival, emphasizing its role in survivability, meiotic progression, and DNA damage checkpoint-mediated apoptosis. This study reveals complex interactions between herbal extracts and C. elegans processes, shedding light on potential antitumor effects and mechanisms. The findings provide insights into the complex landscape of herbal medicine's impact on a model organism, offering implications for broader applications.

This article provides a general overview of the bio-inspired computing method called DNA-Based Computing (DBC), including its theory and applications. The main theme of DBC is the central dogma of molecular biology (once information of DNA/RNA has got into a protein, it can’t get out again), i.e., DNA to RNA (sequences of four types of nucleotides) and DNA/RNA to protein (sequence of twenty types of amino acids) are allowed, not the reverse ones. Thus, DBC transfers few-element information (DNA/RAN-like) to many-element information (protein-like), solving a given cognitive problem. DBC can take many forms; this article elucidates two main forms, denoted as DBC-1 and DBC-2. Using arbitrary numerical examples, it is shown that DBC-1 can solve the following cognitive problems: “similarity indexing between seemingly different but inherently identical objects” and “recognizing regions of an image separated by a complex boundary.” DBC-2 can solve the following cognitive problem: “pattern recognition when the relevant information is insufficient.” Smart manufacturing-based systems (digital twins and big data analytics) must solve the abovementioned problems to make the manufacturing enablers (machine tools and monitoring systems) more self-reliant and autonomous. Consequently, DBC can improve the cognitive problem-solving ability of smart manufacturing-relevant systems and, thereby, can enhance its biologicalization.

The ubiquitin proteasome system (UPS) utilizes an orchestrated enzymatic cascade of E1, E2, and E3 ligases to add single or multiple ubiquitin-like molecules as post-translational modification (PTM) to proteins. Ubiquitination can alter protein functions and/ or marks ubiquitinated proteins for proteasomal degradation but deubiquitinases (DUBs) can reverse protein ubiquitination. While the importance of DUBs as regulatory factors in the UPS is undisputed, many questions remain on DUB selectivity for protein targeting, their mechanism of action, and the impact of DUBs on the regulation of diverse biological processes. Furthermore, little is known about the expression and role of DUBs in tumors of the human central nervous system (CNS). In this comprehensive review, we have used publicly available transcriptional datasets to determine the gene expression profiles of 99 deubiquitinases (DUBs) from five major DUB families in seven primary pediatric and adult CNS tumor entities. Our analysis identified selected DUBs as potential new functional players and biomarkers with prognostic value in specific subtypes of primary CNS tumors. Collectively, our analysis highlights an emerging role for DUBs in regulating CNS tumor cell biology and offers a rationale for future therapeutic targeting of DUBs in CNS tumors.

The approach based on molecular modeling was developed to study dNTP derivatives characterized by new polymerase-specific properties. For this purpose, the relative efficiency of PCR amplification with modified dUTPs was studied using Taq, Tth, Pfu, Vent, Deep Vent, Vent (exo-), and Deep Vent (exo-) DNA polymerases. The efficiency of PCR amplification with modified dUTPs was compared with the results of molecular modeling using the known 3D structures of KlenTaq polymerase–DNA–dNTP complexes. The dUTPs were C5-modified with bulky functional groups (the Cy5 dye analogs) or lighter aromatic groups. Comparing the experimental data and the results of molecular modeling revealed the decrease of PCR efficiency in the presence of modified dUTPs with an increase in the number of non-covalent bonds between the substituents and the DNA polymerase (about 15% decrease per one extra non-covalent bond). Generalization of the revealed patterns to all the studied polymerases of the A and B families is discussed herein. The number of non-covalent bonds between the substituents and polymerase amino acid residues is proposed to be a potentially variable parameter for regulating enzyme activity.

Susceptibility to photoimmune suppression and photocarcinogenesis is greater in male than in female humans and mice and is exacerbated in female estrogen receptor-beta knockout (ER-β-/-) mice. We previously reported that the active vitamin D hormone, 1,25-dihydroxyvitamin D3 (1,25(OH)2D) applied topically protects against ultraviolet radiation (UV)-induction of cutaneous cyclobutane pyrimidine dimers (CPDs) and suppression of contact hypersensitivity (CHS) in female mice. Here we compare these responses in female versus male Skh:hr1 mice, in ER-β-/- versus wild type C57BL/6 mice, and in female ER-blockaded Skh:hr1 mice. Induction of CPDs was significantly greater in male than female Skh:hr1 mice and was more effectively reduced by 1,25(OH)2D in female Skh:hr1 and C57BL/6 mice, than in male Skh:hr1 or ER-β-/- mice respectively. This correlated with reduced sunburn inflammation by 1,25(OH)2D in female but not male Skh:hr1 mice. Furthermore, although 1,25(OH)2D alone dose-dependently suppressed basal CHS responses in male Skh:hr1 and ER-β-/- mice, UV-induced immunosuppression was universally observed. In female Skh:hr1 and C57BL/6 mice, the immunosuppression was decreased by 1,25(OH)2D dose-dependently, but not in male Skh:hr1, ER-β-/- or ER-blockaded mice. These results reveal a sex bias in genetic, inflammatory and immune photoprotection by 1,25(OH)2D favoring female mice, that is dependent on the presence of ER-β.

KLHL genes and the related KBTBD genes encode components of the Cullin-E3 ubiquitin ligase complex and typically target tissue-specific proteins for degradation, thereby affecting differentiation, homeostasis, metabolism, cell signaling, and the oxidative stress response. Despite their importance in normal cell function and in disease (KLHL40, KLHL41, KBTBD13, KEAP1, and ENC1), previous studies that examined epigenetic factors affecting transcription were predominantly limited to promoter DNA methylation. Using diverse tissue and cell culture whole-genome profiles, we examined 17 KLHL or KBTBD genes preferentially expressed in skeletal muscle or brain to identify tissue-specific enhancer and promoter chromatin, open chromatin (DNaseI hypersensitivity), and DNA hypomethylation. Sixteen of the 17 genes displayed muscle- or brain-specific enhancer chromatin in their gene bodies, and most exhibited specific intergenic enhancer chromatin as well. Seven genes were embedded in a super-enhancer. The enhancer chromatin regions typically displayed foci of DNA hypomethylation at peaks of open chromatin. In addition, we found evidence that gene neighbors (HHATL and FBXO32) of KLHL40 and KLHL38 harbor enhancer chromatin that likely upregulates the adjacent KLHL gene. Many KLHL/KBTBD genes had tissue-specific promoter chromatin at their 5’ ends, but surprisingly, two (KBTBD11 and KLHL31) had constitutively unmethylated promoter chromatin in their 3’ exons that overlaps a retrotransposed KLHL gene. Our findings demonstrate the importance of expanding epigenetic analyses beyond the 5’ ends of genes in studies of normal and abnormal gene regulation.

Mitochondrial malfunction is supposed to be involved in the etiology and pathology of major depressive disorder (MDD). Here, we aimed to identify and characterize the molecular pathomechanisms related to mitochondrial disfunction in adult human skin fibroblasts which were derived from MDD patients or non-depressive control subjects. We found that MDD fibroblasts showed significantly impaired mitochondrial functioning: basal and maximal respiration, spare respiratory capacity, non-mitochondrial respiration and ATP-related oxygen consumption was lower. Moreover, MDD fibroblasts harbor lower ATP levels and showed hyperpolarized mitochondrial membrane potential. To investigate cellular resilience, we challenged both groups of fibroblasts with hormonal (dexamethasone) or metabolic (galactose) stress for one week, and found that both stressors increased oxygen consumption but lowered ATP content in MDD as well as in non-depressive control fibroblasts. Interestingly, the bioenergetic differences between fibroblasts from MDD or non-depressed subjects, which were observed under non-treated conditions, could not be detected after stress. Our findings support the hypothesis that altered mitochondrial function causes a bioenergetic imbalance which is associated with the molecular pathophysiology of MDD. The observed alterations in OXPHOS and other mitochondria-related properties represent a basis for further investigations of pathophysiological mechanisms and might open new ways to gain insight into antidepressant signaling pathways.

Due to the rapid developments in aquaculture industry, Artemia franciscana, originally an American species, has been intentionally introduced to the Eurasia, Africa and Australia. In the present study, we used a partial sequence of the mitochondrial DNA Cytochrome Oxidase subunit I (mt-DNA COI) gene and genomic fingerprinting by Inter-Simple Sequence Repeats (ISSRs) to determine the genetic variability and population structure of Artemia populations (indigenous and introduced) from 14 different geographical locations in Western Asia. Based on the haplotype spanning network, Artemia urmiana has exhibited higher genetic variation than native parthenogenetic populations. Although A. urmiana represented a completely private haplotype distribution, no apparent genetic structure was recognized among the native parthenogenetic and invasive A. franciscana populations. Our ISSR findings have documented that despite invasive populations have lower variation than source population in Great Salt Lake (Utah, USA), they have significantly revealed higher genetic variability compare to the native populations in Western Asia. According to the ISSR results, the native populations were not fully differentiated by the PCoA analysis, but the exotic A. franciscana populations were geographically divided in four genetic groups. We believe that during the colonization, invasive populations have experienced substantial genetic divergences, under new ecological conditions in the non-indigenous regions.

Ovarian cancer (OC) represents a significant challenge in the realm of gynecological cancers, characterized by poor survival rates and complex etiology. In this review, we delve into the se-lected genetic, environmental, and hormonal factors underpinning OC development. We have reviewed scientific databases searching for ovarian cancer genetic and epigenetic factors. We have included studies based on their relevancy. As a result of exploring ovarian carcinogen-esis, this systematic review contains data collected from 126 various works. The role of prominent genetic players such as BRCA mutations, DNA repair mechanisms, un-derscores the intricate landscape of OC susceptibility. We explore Li-Fraumeni and Lynch syn-drome, which impart a heightened predisposition to OC development. Hormonal factors such as estrogen, progesterone, and androgens are also discussed. Environmental alterations, ranging from lifestyle influences on OC to microbiome dysbiosis and lifestyle factors such as obesity, al-cohol consumption, and physical activity intersect with genetic and epigenetic pathways, shaping the risk landscape for OC. Through a meticulous examination of current literature, this review provides a nuanced under-standing of the multifactorial nature of OC, emphasizing the need for holistic approaches to prevention, diagnosis, and treatment.

Background: The rise of new SARS-CoV-2 mutations brought challenges and progress in the global fight against COVID-19. Mutations in spike and accessory genes affect transmission, vaccine efficacy, treatments, testing, and public health strategies. Monitoring emerging variants is crucial to halt virus spread. Methods: 44 nasopharyngeal/oropharyngeal swabs from Kenyan patients were sequenced with the Illumina platform. Galaxy's bioinformatic tools were used for genomic analysis. SARS-CoV-2 genome classification was done using PANGOLIN and mutation annotation with the COVID-19 Annotator tool. Results: The study showed 5 clades to be circulating in the region. 38(86%) were BA.1.1; 2(5%) were BA.1.1.1; 1(2%) was BA.1; 1(2%) was BA.1.14 and 2(5%) were AY.46. These clades had a cumulative of 173 mutations among them with 50 novel mutations. Forty-eight of these novel mutations occurred in a low frequency of 2.3% of the sequences tested while the other two, S:R214R, and NSP2:A555A, were for 43.2% and 18.2% of the cases respectively. Conclusions: The high-frequency novel mutations were synonymous mutations, a phenomenon that was previously viewed as phenotypically silent but recent studies indicate they can affect viral fitness with potential functional associations. These findings add to the understanding of the SARS-CoV-2 virus future evolutionary and immunological dynamics in the region

Leukemia encompasses diverse blood cancers marked by the unchecked growth and developmental blockage of myeloid or lymphoid precursor cells within the bone marrow and the bloodstream. Treatment traditionally involves intensive chemotherapy or hematopoietic stem cell transplantation, albeit these approaches often yield severe side effects and encounter resistance from leukemia cells. Hence, the quest for novel therapies targeting leukemic cells selectively is paramount to enhancing leukemia outcomes. Exploring natural products is a promising avenue for pursuing effective chemotherapy and preventive measures against leukemia. These compounds serve as a crucial wellspring for drug development and offer a source of biologically active elements for potential therapeutic interventions in leukemia. The enzyme telomerase plays a vital role in stabilizing chromosomes by adding DNA sections to the end of chromosomes during the mitotic cycle. Notably, hTERT mRNA expression strongly correlates with telomerase activity, making it a potential target for therapeutic intervention without adverse systemic effects. Telomerase inhibition is a promising target in cancer therapy, especially in combination with natural ingredients, demonstrating less cytotoxicity than chemotherapies.

Background: Glioblastoma remains a challenging brain tumor to treat due to its infiltrative nature. Accurately identifying tumor boundaries during surgery is crucial for successful tumor resection. This study introduces an innovative intraoperative visualization method utilizing a surgical fluorescence microscope to precisely locate tumor cells. The focus of the study is on IR-Glint, a novel preparation comprising Cy7.5-labeled aptamers Gli-233nt and Gli-55_3L specific to human glioblastoma. Methods: The aptamers were modified using molecular modeling and quantum chemical techniques. The effectiveness of the preparation was assessed using flow cytometry and microscopy on primary cultures of human glioblastoma. In vivo studies were conducted on mouse and rabbit models, employing orthotopic xenotransplantation of human brain glioblastoma with various imaging techniques, including PET/CT, in vivo fluorescence visualization, confocal laser scanning, and surgical microscopy. Results: The experiments validated the potential of IR-Glint for intraoperative visualization of glioblastoma using infrared imaging. Surface application of the aptamer-based formulation on the brain allowed clear visualization of the tumor, aiding surgeons in tumor resection. This approach also reduces the dosage required and mitigates potential toxic effects on the patient. Conclusions: This study demonstrates the promising potential of using in-frared dye-labeled aptamers for intraoperative visualization of glioblastoma. By improving surgical treatment outcomes in neurosurgery, this novel approach may pave the way for future advancements in the field.

Advanced paternal age at fertilization has been suggested to be a risk factor for neurodevelopmental, psychiatric and other disorders in offspring. One emerging hypothesis suggests that altered offspring phenotype is linked with age-related accumulation of epigenetic changes in the sperm of fathers. Given that paternal age is increasing in the developed world, understanding aging-related epigenetic changes in sperm is needed as well as environmental factors that modify such changes. In this study, we characterize age-dependent changes in sperm DNA methylation profiles between young pubertal (postnatal day (PNDs) 65) and mature (PND120) Wistar rats. We also analyze these changes in rats exposed perinatally to 0.2 mg/kg of ubiquitous environmental xenobiotic 2,2’,4,4’-tetrabromodiphenyl ether (BDE-47). Reduced representation bisulfite sequencing (RRBS) libraries were prepared from caudal epididymal sperm DNA and differentially methylated regions (DMRs; ≥ 10x coverage depth, ≥ 3 CpGs per cluster, ≥ 5% methylation change, q < 0.05) were identified via MethPipe package. In control animals, 5,319 age-dependent DMRs were identified, with 99.3% DMRs hypermethylated in mature animals compared to young pubertal rats. These age-related DMRs were enriched for functional categories essential for embryonic development, such as pattern specification, forebrain and sensory organ development, Hippo and Wnt pathways. Age-related changes in sncRNA, reported in different study, target similar list of genes and biological categories.In BDE-47 exposed rats, sperm DNA methylation was higher in young pubertal and lower in mature animals when compared to controls, which resulted in a significant attenuation in the number of age-dependent DMRs (N = 189) identified in the exposed group. In conclusion, our results indicate that the natural aging process has profound effects on sperm methylation levels and this effect may be modified by environmental exposures. Moreover, our results further support the role of epigenetic mechanisms as a likely link betwen paternal age and offspring health and development.

RNA polymerase II (RNAPII) frequently transcribes non-protein coding DNA sequences in eukaryotic genomes into long non-coding RNA (lncRNA). Here, we focus on the impact of the act of lncRNA transcription on nearby functional DNA units. Distinct molecular mechanisms linked to the position of lncRNA relative to the coding gene illustrate how non-coding transcription controls gene expression. We review the biological significance of the act of lncRNA transcription on DNA processing, highlighting common themes, such as mediating cellular responses to environmental changes. This review presents the background in chromatin signaling to appreciate examples in different organisms where we can interpret functions of non-coding DNA through the act of RNAPII transcription.

Diffuse intrinsic pontine glioma (DIPG) is an aggressive pediatric brainstem tumor with a 5-year survival of <1%. Up to 80% of DIPG tumors contain a specific K27M mutation in one of two genes encoding histone H3 (H3K27M). Furthermore, p53 mutations found in >70-80% of H3K27M DIPG, and mutant p53 status is associated with a decreased response to radiation treatment and worse overall prognosis. Recent evidence indicates that H3K27M mutation disrupts tri-methylation at H3K27 leading to aberrant gene expression. Jumonji family histone demethylases collaborates with H3K27 mutation in DIPG by erasing H3K27 trimethylation and thus contributing to derepression of genes involved in tumorigenesis. Since the first line treatment for pediatric DIPG is fractionated radiation, we investigated the effects of Jumonji demethylase inhibition with GSK-J4, and mutant p53 targeting/oxidative stress induction with APR-246, on radio-sensitization of human H3K27M DIPG cells. Both APR-246 and GSK-J4 displayed growth inhibitory effects as single agents in H3K27M DIPG cells. Furthermore, both of these agents elicited mild radiosensitizing effects in human DIPG cells (sensitizer enhancement ratios (SERs) of 1.12 and 1.35, respectively; p<0.05). Strikingly, a combination of APR-246 and GSK-J4 displayed a significant enhancement of radiosensitization, with SER of 1.50 (p<0.05) at sub-micro-molar concentrations of the drugs (0.5 μM). The molecular mechanism of the observed radiosensitization appears to involve DNA damage repair deficiency triggered by APR-246/GSK-J4, leading to the induction of apoptotic cell death. Thus, a therapeutic approach of combined targeting of mutant p53, oxidative stress induction, and Jumonji demethylase inhibition with radiation in DIPG warrants further investigation.

We propose an improved earlier described “mirror” method [1] for detecting in cell nuclear extracts mutations that arise in DNA during its replication due to misincorporation of deoxyadenosine-5’-monophosphate (dAMP) opposite 7,8-dihydro-8-oxoguanine (8-oxoG). The method is based on the synthesis of a complementary chain (“mirror”) by nuclear extracts of different mice organs on a template containing 8-oxoG inside and dideoxycytidine residue (ddC) at the 3’-end. The “mirror”was amplified by PCR using primers part of which was non-complementary to the template. It allowed obtaining the “framed mirror” products. The misincorporation of dAMP in “framed mirror” products forms an EcoRI restriction site. The restriction analysis of double-stranded “framed mirror” products allows a quantification of the mutation frequency in nuclear extracts. The data obtained showed that the mutagenic potential of 8-oxoG markedly varied in different organs of adult mice and embryos.

DNA methylation and other epigenetic factors are important in the pathogenesis of late-onset Alzheimer’s disease (LOAD). Methylenetetrahydrofolate reductase (MTHFR) gene mutations occur in most elderly patients with memory loss. MTHFR is critical for production of S-adenosyl-L-methionine (SAM), the principal methyl donor. A common mutation (1364T/T) of the cystathionine-γ-lyase (CTH) gene affects the enzyme that converts cystathionine to cysteine in the trans-sulfuration pathway causing plasma elevation of total homocysteine (tHcy) or hyperhomocysteinemia – a strong and independent risk factor for cognitive loss and AD. Other causes of hyperhomocysteinemia include aging, nutritional factors, and deficiencies of B vitamins. We emphasize the importance of supplementing vitamin B₁₂ (methylcobalamin), vitamin B₉ (folic acid), vitamin B₆ (pyridoxine), and SAM to patients in early stages of LOAD.

Plants thriving in desert environments are suitable for studying mechanisms for plant survival under extreme seasonal climate variation. Zygophyllum dumosum Boiss, like many other Zygophyllaceae species, displays a unique epigenetic mechanism whereby the repressive markers di- and tri-methyl of H3K9 do not exist. We studied epigenetic mechanisms underlying seasonal growth cycles in Z. dumosum and their association with factors regulating basic cell functions. We showed strong association between rainfall and seasonal growth and the epigenetic marker of dimethyl H3K4, which disappears on entry into the dry season and the acquisition of dormant state. DNA methylation is not affected by lack of H3K9 di and tri methyl and changes in methylation pattern are apparent on entry into the dry season. Proteome analysis of acid soluble fractions revealed extensive reduction in ribosomal proteins and in proteins involved in chloroplasts and mitochondria activities during the dry seasons concomitantly with up-regulation of molecular chaperone HSPs. Our results highlight mechanisms underlying Z. dumosum adaptation to seasonal climate variation. Particularly, summer dormancy is associated with loss of the permissive epigenetic marker dimethyl H3K4, which might facilitate genome compaction, concomitantly with significant reduction in proteins involved in basic cell functions (i.e., protein synthesis, photosynthesis and respiration).

This article delves into the historical development and ramifications of divergent theories that have emerged since the publication of Darwin's seminal work, "On the Origin of Species" in 1856. It traces the evolution of thought from Darwin's revolutionary ideas through the controversial era of eugenics, the groundbreaking discovery of DNA, the elucidation of Mendelian genetics, and the subsequent development of modern genetic theories. The article also explores the critical role of mathematical models, statistics, and data science in advancing genomics research over the last decade. This comprehensive exploration highlights how these theories and methods have fundamentally shaped our understanding of biology, evolution, human genetics, and complex biological systems.

This study explores the role of inflammation and oxidative stress, hallmarks of COVID-19, in accelerating cellular biological aging. We investigated early molecular markers—DNA methylation age (DNAmAge) and telomere length (TL)—in blood leukocytes, nasal cells (NC), and induced sputum (IS) one-year post-infection in pauci- and asymptomatic healthcare workers (HCWs) infected during the first pandemic wave (February-May 2020), compared to COPD patients, model for "aged lung". Data by questionnaires, Work Ability Index (WAI), blood analyses, autonomic cardiac balance assessments, heart rate variability (HRV), and pulmonary function tests were collected. Elevated leukocyte DNAmAge significantly correlated with advancing age, male gender, daytime work, and an aged phenotype characterized by chronic diseases, elevated LDL and glycemia levels, medications affecting HRV, and declines in lung function, WAI, lymphocyte count, hemoglobin levels, and HRV (p&lt;0.05). Increasing age, LDL levels, job positions involving intensive patient contact, and higher leukocyte counts collectively contributed to shortened leukocyte TL (p&lt;0.05). Notably, HCWs exhibited accelerated biological aging in IS cells compared to both blood leukocytes (p≤0.05) and NC (p&lt;0.001), and were biologically older than COPD patients (p&lt;0.05). These findings suggest the need for monitoring aging in pauci- and asymptomatic COVID-19 survivors, who represent the majority of the general population.

The evidential value of an mtDNA match between biological remains and its potential donor is determined by the random match probability of the haplotype. This probability is based on the haplotype’s population frequency estimate. Consequently, implementing a population study representative of the population relevant to the forensic case is vital to correctly evaluating the evidence. The emerging number of poaching cases and the limited availability of such data emphasizes the need for improved fallow deer mtDNA population databank for forensic purposes, including targeting the entire mitochondrial control region. By sequencing the whole mitochondrial control region in 138 animals from five populations in Hungary we found four different haplotypes, including one which has not yet been described. Our results, supplemented with data already available from previous research do not support the possibility of determining the population of origin, although some patterns of geographical separation can be distinguished. Estimates of molecular diversity indicate approximately similarly low mtDNA diversity (Hd=0.565 and π = 0.002), compared with data from other countries. The calculated random match probability of 0.547 shows a high probability of coincidence and, therefore, a limited capacity for exclusion. Consequently, it may be necessary to search for polymorphic sites on other mitochondrial gene sections or to sequence the entire mitogenome of fallow deer.

Varunaivibrio sulfuroxidans type strain TC8T is a mesophilic, facultatively anaerobic, facultatively chemolithoautotrophic alphaproteobacterium, isolated from a sulfidic shallow-water marine gas vent located at Tor Caldara, Tyrrhenian Sea, Italy. V. sulfuroxidans belongs to the family Thalassospiraceae within the Alphaproteobacteria, with Magnetovibrio blakemorei as its closest relative. The genome of V. sulfuroxidans encodes the genes involved in sulfur, thiosulfate and sulfide oxidation, as well as nitrate and oxygen respiration. The genome encodes the genes involved in carbon fixation via Calvin-Benson-Bassham cycle, in addition to genes involved in glycolysis and TCA cycle, indicating a mixotrophic lifestyle. Genes involved in the detoxification of mercury and arsenate are also present. The genome also encodes a complete flagellar complex, one intact prophage and one CRISPR, as well as a putative DNA uptake mechanism mediated by the type IVc (aka Tad pilus) secretion system. Overall, the genome of Varunaivibrio sulfuroxidans indicates that the bacterium is metabolically versatile and well-adapted to the dynamic environmental conditions of sulfidic gas vents.

Apolipoprotein B mRNA-editing enzyme catalytic polypeptide-like (APOBEC) proteins are a diverse and evolutionarily conserved family of cytidine deaminases that provide a variety of functions from tissue-specific gene expression and immunoglobulin diversity to control of viruses and retrotransposons. APOBEC family expansion has been documented among mammalian species, suggesting a powerful selection for their activity. Enzymes with a duplicated zinc-binding domain often have catalytically active and inactive domains, yet both have antiviral function. Although APOBEC antiviral function was discovered through hypermutation of HIV-1 genomes lacking an active Vif protein, much evidence indicates that APOBECs also inhibit virus replication through mechanisms other than mutagenesis. Multiple steps of the viral replication cycle may be affected, although nucleic acid replication is a primary target. Packaging of APOBECs into virions was first noted with HIV-1, yet is not a prerequisite for viral inhibition. APOBEC antagonism may occur in viral producer and recipient cells. Signatures of APOBEC activity include G-to-A and C-to-T mutations in a particular sequence context. The importance of APOBEC activity for viral inhibition is reflected in the identification of numerous viral factors, including Vif, which are dedicated to antagonism of these deaminases. Such viral antagonists often are only partially successful, leading to selection for viral variants.

The study presents a mathematical modeling for supporting the premise that our body is composed of blockchain systems. A cell population is considered a space with peer-to-peer communication networks for applying blockchain protocol. Transaction is defined as gene expression that constantly occurs for protein synthesis in each cell. The transaction process proceeds according to the blockchain protocol with sharing and recording the data on the blockchain ledger. The premise comes from the clues of the previous research such that the cell population is a complex structured network system having cell-to-cell connections. Although individual cells exhibit stochastic nonlinear dynamic behavior, cell population shows consensus behavior that reaches to ensemble through interaction among cells. It is inferred that gene expression is not regulated by the corresponding cell only nor determined by external intelligence such as the brain. This is achieved through a consensus through stochastic interactions between cells over the whole cell population. These findings imply that mathematical blockchain modeling is a suitable for gene expression process. The original contribution of the study is a methodology for applying mathematically the blockchain protocol to the real biological gene expression process. In other words, the DNA sequence is converted into a numeral bit sequence that makes it possible to implement the blockchain protocol. A new DNA sequence scheme is proposed with adding methylated cytosine and adenine as the 5th and 6th bases for including epigenetic information which has profound effect on gene expression and regulation. The methodology was applied to the real biological synthesis process of protein samples. The protein is composed of amino acids that are encoded by triplet codons of 216 kinds with 6 base RNA sequence. The gene expression information is traced backward from a synthesized protein sample, amino acids of codons, RNA transcript up to DNA sequence. One of the results is a numeric value in the form of a bit sequence with which mathematical blockchain modeling is applicable. The cryptographic authentication and the consensus process are mathematically proven to work properly by the blockchain protocol. It implies that the same protein is synthesized, but with different epigenetic data, then protein's latent material properties will certainly be different. Although the result has not been justified by the biologic experiment yet, it is sure that the biological hidden algorithm inside DNA sequence will be revealed by the binary bit-logic with physical on/off states which is mathematically proven. The research will greatly contribute to disease treatment and medicine development as well as epigenetics in the future.

The extracellular matrix of most bacterial biofilms contains polysaccharides, proteins, and nucleic acids. These biopolymers have been shown to mediate fundamental biofilm-related phenotypes including surface attachment, intercellular adhesion, and biocide resistance. Enzymes that degrade polymeric biofilm matrix components, including glycoside hydrolases, proteases, and nucleases, are useful tools for studying the structure and function of biofilm matrix components and are also being investigated as potential antibiofilm agents for clinical use. Dispersin B is a well-studied, broad spectrum antibiofilm glycoside hydrolase produced by Aggregatibacter actinomycetemcomitans. Dispersin B degrades poly-N-acetylglucosamine, a biofilm matrix polysaccharide that mediates biofilm formation, stress tolerance, and biocide resistance in numerous Gram-negative and Gram-positive pathogens. Dispersin B has been shown to inhibit biofilm and pellicle formation; detach pre-formed biofilms; disaggregate bacterial flocs; sensitize pre-formed biofilms to detachment by enzymes, detergents, and metal chelators; and sensitize pre-formed biofilms to killing by antiseptics, antibiotics, bacteriophages, macrophages, and predatory bacteria. This review summarizes the results of nearly 100 in vitro and in vivo studies that have been carried out on dispersin B since it’s discovery 20 years ago. These include investigations into the biological function of the enzyme, it’s structure and mechanism of action, and its in vitro and in vivo antibiofilm activities against numerous bacterial species. Also discussed are potential clinical applications of dispersin B.

According to many research groups, high glucose induces over production of superoxide anions, and reactive oxygen species (ROS) in general is the link between high glucose levels and the toxicities seen at cellular levels. Respiratory complex anomalies could lead to the production of ROS. Calcium [Ca2+], at physiological levels serves as a second messenger in many physiological functions. Accordingly, mitochondrial calcium [Ca2+]m overload leads to ROS production which could be lethal to the mitochondria through various mechanisms. The F1F0-ATPase (ATP syn-thase or complex V) is the enzyme that is responsible for catalyzing the final step of oxidative phosphorylation. It does so by coupling the translocation of protons in the mitochondrial in-termembrane space and shuttling them to the mitochondrial matrix for ATP synthesis to take place. Mitochondrial complex V T8993G mutation specifically blocks the translocation of protons across the intermembrane space thereby blocking ATP synthesis leading to Neuropathy, Ataxia and Retinitis Pigmentosa Syndromes (NARP). This study seeks to explore the possibility of [Ca2+]m overload mediating the pathological roles of high glucose in defective respiratory chain mediated mitochondrial stress. The NARP cybrids are the in vitro experimental models for the cells with F1FO-ATPase defects, these cells harbor 98% of mtDNA T8993G mutation. Their coun-terparts 143B osteosarcoma cell lines are the parental cell lines used for comparisons. We uncov-ered that, NARP cells mediated and enhanced the death of the cells (apoptosis) when incubated with hydrogen peroxide (H2O2) and high glucose respectively as depicted by the MTT assay of cell viability. Furthermore, using the fluorescence probe-coupled laser scanning confocal imag-ing microscopy, NARP cells significantly enabled mitochondrial reactive oxygen species (mROS) formation and enhanced the depolarization of the mitochondrial membrane potential (ΔΨm). Elucidating the mechanisms of sugar enhanced toxicity on mitochondria would help in the fu-ture to alleviate the symptoms of patients associated with (NARP) syndromes.

(1) Background: Double-strand breaks (DSBs) in a single nucleus are usually measured using the sperm chromatin structure assay (SCSA), sperm chromatin dispersion (SCD) test, and comet assay (CA). We developed mono-dimensional single-cell pulsed-field gel electrophoresis (1D-SCPFGE) and angle-modulated two-dimensional single-cell pulsed-field gel electrophoresis (2D-SCPFGE) to observe DNA fragmentation in separated motile sperm. (2) Methods: We set up comparative standards, calibration curves, the required sensitivity, and eligibility criteria for test sperm to validate their measurement principles. (3) Results: We revealed that the conventional methods overlooked the interference of nucleoproteins in their measurements. In-gel proteolysis improves the measurement accuracies of 1D- and 2D-SCPFGE. We recommend using naked DNA for comparative standards and test specimens. Moreover, we have observed several dysfunctions that may induce DNA damage in the separated motile sperm. Overall, our discussion highlights the need to revisit the conventional univariable analyses based on the SCSA, SCD test, and CA. (4) Conclusions: Human infertility is a complex syndrome, and the aim of quality control in intracytoplasmic sperm injection is to identify the underlying dysfunctions remaining in the separated motile sperm that render them ineligible for injection. Well-designed, multivariable analyses with special consideration to confounding factors are necessary in future cohort studies.

Various attempts to amplify an AQP11 cDNA from tissues of the spiny dogfish (Squalus acanthias) were made. Two pairs of deoxy-inosine-containing degenerate primers were designed based on conserved amino acid sequences from an AQP11 alignment. These primers yielded some faint bands from gill cDNA that were sequenced. Blast searches with the sequences showed they were not AQP11. An elasmobranch AQP11 nucleotide sequence alignment was produced to identify conserved regions to make further degenerate primers. One primer pair produced a short 148bp fragment showing particularly strong expression in gill and intestine. It was sequenced and represented a piece of the AQP11 gene. However, as the fragment may have resulted from contaminating genomic DNA (in total RNA used to make cDNA), 5’ and 3’ RACE was performed to amplify the two ends of the putative cDNA. 5’ and 3’ RACE amplifications depend on the presence of a 5’ cap nucleotide and a poly A tail respectively on the putative AQP11 mRNA. Hence successful amplification was only possible from cDNA and not genomic DNA. Nested RACE amplifications were performed using gill and intestinal RACE cDNA but none of the DNA fragments were AQP11. Consequently, the spiny dogfish AQP11 gene may represent a pseudogene.

Fanconi anemia (FA) develops due to a mutation in one of the FANC genes that are involved in repair of interstrand crosslinks (ICLs). FANCG, a member of the FA core complex, is essential for ICL repair. Previous FANCG-deficient mouse models were generated with drug-based selection cassettes in mixed mice backgrounds, leading to a disparity in interpretation of genotype-related phenotype. To exclude these confounders, we created a Fancg-KO (KO) mouse model using CRISPR/Cas9. The entire Fancg locus was targeted and maintained on the immunological well-characterized C57BL/6J background. Intercrossing of heterozygous mice resulted in sub-Mendelian numbers of homozygous mice, suggesting loss of FANCG can be embryonic lethal. KO mice displayed infertility, hypogonadism but no other developmental problems. Bone marrow analysis revealed a defect in various hematopoietic stem and progenitor subsets with a bias towards myelopoiesis. Cell lines derived from Fancg-KO mice were hypersensitive to crosslinking agents-cisplatin and Mitomycin C, and Fancg-KO mouse embryonic fibroblasts (MEFs) displayed increased γ-H2AX upon cisplatin treatment. Reconstitution of these MEFs with Fancg cDNA corrected for the ICL hypersensitivity. Collectively, this project provides a new, genetically and immunologically well-defined Fancg-KO mouse model for further in vivo and in vitro studies on FANCG and ICL repair.

ORF6 is responsible for suppressing the immune response of cells infected by the SARS-CoV-2 virus. It is also the most toxic protein of SARS-CoV-2, and its actions are associated with the viral pathogenicity. Here we study in silico and in vitro the structure of the protein, its interaction with RAE1, and the mechanism of action behind its high toxicity. We show both computationally and experimentally that SARS-CoV-2 ORF6, embedded in the membranes of the endoplasmic reticulum, binds to RAE1 and sequesters it in the cytoplasm, thus depleting its availability in the nucleus and impairing nucleocytoplasmic mRNA transport. This negatively affects the cellular genome stability by compromising the cell cycle progression into the S-phase and by promoting the accumulation of RNA-DNA hybrids. Understanding the multiple ways in which ORF6 affects DNA replication may also have important implications for elucidating the pathogenesis of SARS-CoV-2 and developing therapeutic strategies to mitigate its deleterious effects on host cells.

This survey reports on the DNA identification and occurrence of Culex torrentium and Cx. pipiens s.s. in Belgium. These native disease vector mosquito species are morphologically difficult to separate and the biotypes of Cx. pipiens s.s. are morphologically indistinguishable. Culex torrentium and Cx. pipiens s.s. were identified using the COI and ACE2 loci. We recorded 1,248 Cx. pipiens s.s. and 401 Cx. torrentium specimens from 24 locations in Belgium (collected between 2017 and 2019). Culex pipiens biotypes pipiens and molestus, and their hybrids, were differentiated by fragment size analysis of the CQ11 locus (956 pipiens and 227 molestus biotype specimens, 29 hybrids). Hybrids were observed at 13 out of 16 sympatric sites. These results confirm that both species are widespread in Belgium, but while Cx. torrentium revealed many COI haplotypes, Cx. pipiens s.s. showed only one abundant haplotype. This latter observation may either reflect a recent population-wide demographic or range expansion, or a recent bottleneck, possibly linked to a Wolbachia infection. Finally, new evidence is provided for the asymmetric but limited introgression of the molestus biotype into the pipiens biotype.

Chemotherapy represents the most applied approach to cancer treatment. Owing to the frequent onset of chemoresistance and tumor relapses, there is an urgent need to discover novel and more effective anticancer drugs. In search for therapeutic alternatives to treat the cancer disease, a se-ries of hybrid pyrazolo[3,4-d]pyrimidin-4(5H)-ones tethered with hydrazide-hydrazones, 5a-h, was synthesized from condensation reaction of pyrazolopyrimidinone-hydrazide 4 with a series of arylaldehydes in EtOH, in acid catalysis. In vitro assessment of antiproliferative effects against MCF-7 breast cancer cells, unveiled that 5a, 5e, 5g, and 5h were the most effective compounds of the series and exerted their cytotoxic activity through apoptosis induction and G0/G1 phase cell-cycle arrest. To explore their mechanism at a molecular level, 5a, 5e, 5g, and 5h were evalu-ated for their binding interactions with two well-known anticancer targets, namely the epider-mal growth factor receptor (EGFR) and the G-quadruplex DNA structures. Molecular docking simulations highlighted high binding affinity of 5a, 5e, 5g, and 5h towards EGFR. CD experi-ments suggested 5a as a stabilizer agent of the G-quadruplex from the KRAS oncogene promot-er. In the light of these findings, we propose the pyrazolo-pyrimidinone scaffold bearing a hy-drazide-hydrazone moiety as a lead skeleton for designing novel anticancer compounds.

Translation is spatiotemporally regulated and ER-associated mRNAs are generally in efficient translation. It is unclear whether the ER-associated mRNAs are deadenylated or degraded on the ER surface in situ or in the cytosol. Here, we showed that ER possessed active deadenylases, particularly the poly(A)-specific ribonuclease (PARN), in common cell lines and mouse tissues. Consistently, purified recombinant PARN exhibited a strong ability to insert into the Langmuir monolayer and liposome. ER-anchored PARN was found to be able to reshape the poly(A) length profile of the ER-associated RNAs by suppressing long poly(A) tails without significantly influencing the cytosolic RNAs. The shortening of long poly(A) tails did not affect global translation efficiency, suggesting that the non-specific action of PARN towards long poly(A) tails was beyond the scope of translation regulation on the ER surface. Transcriptome sequencing analysis indicated that the ER-anchored PARN trigged the degradation of a small subset of ER-enriched transcripts. The ER-anchored PARN modulated the translation of its targets by redistributing ribosomes to heavy polysomes, suggesting that PARN may play a role in dynamic ribosome reallocation. During DNA damage response, MK2 phosphorylated PARN-Ser557 to modulate PARN translocation from the ER to cytosol. By promoting the decay of ER-associated MDM2 transcripts with low ribosome occupancy, the ER-anchored PARN modulated DNA damage response and thereby cell viability. These findings revealed that a highly regulated communication between mRNA degradation rate and translation efficiency is present on the ER surface in situ and that PARN may contribute to this communication by modulating the dynamic ribosome reallocation between transcripts with low and high ribosome occupancies.

The vaccine industry has undoubtedly contributed massively in the medical world, with hundreds of millions of lives saved as a result of the successful development of prophylactic and therapeutic vaccines against various infectious and oncological diseases. Simultaneously, there has been an ongoing evolutionary battle between polymorphic microbes and the human immune system, and various microbes have developed counter-responses against the human host immune system, by creating diverse “holes” of vulnerability in the innate immune system. Such mechanisms may have resulted due to the lower prophylactic and therapeutic focus upon sharpening the innate immune lines as directly as necessary. The overall objective of vaccine-development should be the sharpening of all immune departments that are targeted by various microbial agents for the purpose of transient innate immune suppression and the distribution of the microbial count/load at a systemic level. Likewise, it could be viable for the clinical research community to start focusing upon filling in the “gaps” of vulnerability that are now present in the pathways responsible for the proper natural immune activation and proportionate signalling toward the adaptive immune system. Perhaps, through such a process, the “road” toward the adaptive immune system, which is “old” and filled with “holes”, could be gradually transformed into a perfectly uniform “highway”, with all the required signals transmitted robustly from the innate to the adaptive immune system, before microbes manage to translate the various pathogenic proteins that transiently prevent such transmission altogether, until it becomes too late for the immune response to possibly develop in a proportionate and non-harmful manner to the organism. The potential “missing piece of puzzle” in vaccine-based clinical research could involve training of the human organism to develop immune memory without the host organism reaching a threshold of clinical disease onset even if viruses translate proteins that silence the host interferon system and make it unable to trace produced toxins whilst not directly causing clinical pathophysiology. Whilst it is realistically almost impossible to train the human immune system to develop responses and memory against viral proteins that do not directly cause immunopathology, there could be a set of clinical approaches involving a stimulation for more robust activation of the host interferon system, with an example of such stimuli being the same principal elements of the interferon system. Another example of pharmaceutical approaches would involve the development of drug-like components to target such viral proteins, though they would be more costly and consequently more restricted in usage; due to the fact that viruses are obligate intracellular, particle-based parasites and can only synthesise their proteins inside host cells. Such a scenario would be possible, given that it is almost impossible for human evolutionary growth to eventually stall and not risk experiencing a decrease as a result. Simultaneously, it is important for human society to bind by the principles of maintaining a healthy and nature-friendly lifestyle, given the philosophical quote of “We are what we eat.” The overall objective should be the facilitation of human and animal immune evolution over viral and bacterial molecular self-camouflaging from all possible directions, perhaps by transferring a concept called “a United Immune System” into clinical practice, as a sharper human and animal evolutionary growth resulting from such actions could “cost” the sharper evolutionary growth of microbes as such, given that evolutionary patterns are based upon the models observed in Physics, in Newton’s Third Law of Motion, as well as in the First Law of Thermodynamics, which concerns the Conservation of Energy.

Background: Existing human cell immortalization methods made the cells obtain oncogenesis phenotype and/or caused the cells gain and/or lose chromosomes. Immortalized normal human T cells lines provide critical in vitro models for basic research and therapeutic products development.Methods: We utilized a CRISPR/Cas9 system to replace a single copy of the exon 2 of the cell cycle inhibitor gene CDKN2A (encoding p16 and p14 proteins) with a single copy of human telomerase reverse transcriptase (hTERT) to immortalize human primary CD8⁺ T cells (hCD8⁺T-TERT). By using Cas9 protein and low donor DNA copies/cell, we successfully immortalized hCD8⁺T cells with a single copy of hTERT transgene, which also avoided uncontrolled insertion of Cas9 gene and guide RNA vector.Results: Human primary CD8+ cells were immortalized and expanded more than 2.6 × 10⁷ times. Characterization of the cells revealed that the immortalized CD8⁺ T-TERT cells retained most of the cell surface markers and normal karyotype. The CD8⁺ T-TERT cells also retained the dependence of IL-2 and CD3/CD28 activator for survival and expansion.Conclusion: we established a stable immortalized CD8⁺ T cell line that had a phenotype consistent with T cells and will be a useful research model.

Barrett’s esophagus (BE) is a known precursor to Esophageal Adenocarcinoma (EAC). Guidelines recommend BE screening in populations with multiple risk factors, for which non-endoscopic esophageal cell collection with biomarker testing is considered an acceptable alternative to esophagogastroduodenoscopy (EGD). This study presents analytical validation of EsoGuard® (EG), a two gene DNA methylation biomarker assay, in esophageal cell samples collected with the 510(k)-cleared, swallowable EsoCheck® (EC) device. EG is a Next Generation Sequencing (NGS) assay that evaluates methylated Vimentin (VIM) and Cyclin A1 (CCNA1) genes, clinically validated biomarkers for detection of BE and EAC. The studies were conducted according to standards of College of American Pathology (CAP), Clinical Laboratory Improvement Amendments (CLIA), and New York (NY) state requirements for analytical validation of molecular assays. Comparison to Sanger sequencing showed EG was 100% accurate at all 31 CpG sites evaluated by the assay. Analytical sensitivity, specificity and accuracy of the assay was 89%, 100% and 96% respectively. Intra- and inter-assay precision was 100%. Limit of detection (LOD) was 1 in 400 methylated cells and reference range was 84%. In summary, EsoGuard is a robust DNA methylation test to detect BE with high accuracy, repeatability, and reproducibility in samples collected using the EsoCheck device.

The exploration of three-dimensional (3D) DNA structures marks a pivotal shift in genomics, offering unprecedented insights into the intricacies of cellular regulation, disease genesis, and the potential for novel diagnostic and therapeutic strategies. This study delves into the dynamic nature of 3D genome organization, investigating how these structures influence cellular functions in response to environmental changes and over time. Through a combination of real-time imaging and advanced computational models, we aim to unravel the complex interplay between 3D DNA structures, non-coding RNAs, and chromatin organization, and their collective impact on gene expression and cellular differentiation. A significant focus is placed on understanding the role of 3D DNA structures in the development and progression of genetic disorders. By employing comprehensive genomic and epigenomic profiling, this research seeks to identify structural aberrations linked to diseases, paving the way for the development of targeted diagnostics and therapies. Additionally, the study explores the potential of genome editing technologies, such as CRISPR-Cas systems, in correcting these structural abnormalities, while also addressing the ethical, legal, and social implications (ELSI) of such interventions. Integrating data from various genomic analyses, we propose the development of advanced computational models to predict the effects of 3D DNA structures on gene expression and cellular function. This integrative approach not only promises to enhance our understanding of genomic regulation but also sets the stage for groundbreaking advances in genomics-based diagnostics and therapeutics. The findings of this study have the potential to revolutionize our approach to treating genetic disorders, offering a new frontier in personalized medicine.

Viruses rely on host cells to replicate their genomes and assemble new viral particles. Thus, they have evolved intricate mechanisms to exploit host factors. Host cells, in turn, have developed strategies to inhibit viruses, resulting in a nuanced interplay of co-evolution between virus and host. This dynamic often involves competition for resources crucial for both host cell survival and virus replication. Iron and iron-containing cofactors, including iron-sulfur clusters, are known to be a heavily battled resource during bacterial infections where control over iron can tug the war in favor of the pathogen or the host. It is logical to assume that viruses also engage in this competition. Surprisingly, our knowledge about how viruses utilize iron (Fe) and iron-sulfur (FeS) clusters remains limited. The handful of reviews on this topic primarily emphasize the significance of iron in supporting the host immune response against viral infections. The aim of this review, however, is to organize our current understanding of how viral proteins utilize FeS clusters, to give perspectives on what questions to ask next and to propose important avenues for future investigations.

For the in-house validation of a droplet digital PCR method, a factorial experimental design was implemented. This design serves different purposes. On the one hand, it is an efficient design in relation to the workload involved in achieving a desirable level of reliability of variance estimates. On the other hand, it allows a partitioning of total variance into different components, thus providing information regarding the dominant sources of random variation. The statistical modelling reflects the actual measurement mechanism, establishing relationships between nominal target DNA copies per well, the range of variation of copy numbers per droplet, probability of detection values, and estimated numbers of copies.

Pericentromeric regions of chromosomes enriched in tandemly repeated satellite DNA although representing a significant part of eukaryotic genomes are still understudied mainly due to interdisciplinary knowledge gaps. Recent studies suggest their important role in genome regulation, karyotype stability and evolution. Thus, the idea of satellite DNA as a junk part of the genome was refuted. Integration of data about molecular composition, chromosome behaviour and details of in situ organization of pericentromeric regions is of great interest. The objective of this work was a cytogenetic analysis of the interactions of pericentromeric regions non-homologous chromosomes in mouse spermatocytes using immuno-FISH. We analysed two events: the associations between cerntomeric regions of X chromosome and autosomes, and associations between centromeric regions of autosomal bivalents forming chromocenters. We conclude that X chromosome form temporary synaptic associations with different autosomes in early meiotic prophase I which normally can be found at pachytene-diplotene without signs of pachytene arrest. These associations are formed between the satellite DNA-enriched centomeric regions of X chromosome and different autosomes but not involve the satellite-poor centromeric region of Y-chromosome. We suggest the mechanism of X chromosome competitive replacement from such associations during synaptic correction. We showed that centromeric region of the X chromosome remains free of γH2Ax-dependent chromatin inactivation, while Y chromosome is completely inactivated. This findings highlights the predominant role of associations between satellite DNA-enriched regions of different chromosomes including X. We assume that X-autosome temporary associations is a manifestation of an additional synaptic disorders checkpoint. These associations are normally corrected before the late diplotene. We revealed that the intense spreading conditions applied to the spermatocytes I nuclei did not lead to destruction of stretched chromatin fibers i.e. elongated chromocenters enriched in satellite DNA. Revealed by us tight associations between pericentromeric regions of different autosomal bivalents and X chromosome may represent the basis for repeat stability maintenance in autosomes an X chromosome. The consequences of our findings are discussed. We obtained the preparations of mouse spermatocytes nuclei in the meiotic prophase I using two approaches: standard and extremely intense surface spread techniques. Using immuno-FISH we visualized tandemly repeated mouse Major and Minor satellite DNA located in the pericentromeric regions of chromosomes and performed a morphological comparison of the standard- and intensely spreaded meiotic nuclei. Based on our results, we assume the remarkable strength of the chromocenter-mediated associations, “chromatin “bridges”, between different bivalents at the pachytene and diplotene stages. We have demonstrated that the chromocenter “bridges” between the centromeric ends of meiotic bivalents are enriched in both tandemly repeated Major and Minor satellite DNA. Association of centromeric regions of autosomal bivalents and X-chromosome but not with Y-chromosome correlates with the absence of Major and Minor satellites on Y-chromosome. We suggest that revealed tight associations between pericentromeric regions of bivalents may represent the network-like system providing dynamic stability of chromosomal territories, as well as add new data for the hypothesis of ectopic recombination in these regions which supports sequence homogeneity between non-homologous chromosomes and does not contradict the meiotic restrictions imposed by the crossing-over interference near centromeres. We conclude that nuclear architecture in meio-sis may play an essential role in contacts between the non-homologous chromosomes providing the specific characteristics of pericentromeric DNA.

Conventional plaque assays rely on the use of overlays to restrict viral infection allowing the formation of distinct foci that grow in time as the replication cycle continues leading to counta-ble plaques that are visualized with standard techniques such as crystal violet, neutral red or immunolabeling. This classical approach takes several days until large enough plaques can be visualized and counted with some variation due to subjectivity in plaque recognition. Since plaques are clonal lesions produced by virus-induced cytopathic effect, we applied DNA fluores-cent dyes with differential cell permeability to visualize them by live cell imaging. We could observe different stages of that cytopathic effect corresponding to an early wave of cells with chromatin-condensation followed by a wave of dead cells with membrane permeabilization within plaques generated by different animal viruses. This approach enables an automated plaque identification using image analysis to increase single plaque resolution compared to crystal violet counterstaining and allows its application to plaque tracking and plaque reduction assays to test compounds for both antiviral and cytotoxic activities. This fluorescent real-time plaque assay sums to those next-generation technologies by combining this robust classical method with modern fluorescence microscopy and image analysis approaches for future applica-tions in virology.

The usage of Quantum Similarity through the equation Z = {∀θ ∈ Z → ∃s ∈ S ∧ ∃t ∈ T : θ = (s, t)}, represents a way to analyze the way communication works in our DNA. Being able to create the object set reference for z being (s, t) in our DNA strands, we are able to set logical tags and representations of our DNA in a completely computational form. This will allow us to have a better understanding of the sequences that happen in our DNA. With this approach, we can also utilize mathematical formulas such as the Euler–Mascheroni constant, regression analysis, and computational proofs to answer important questions on Quantum biology, Quantum similarity, and Theoretical Physics.

Epigenetics, the study of heritable changes in gene expression that do not involve alterations to the DNA sequence, is revolutionizing our understanding of psychiatric disorders. This article provides a comprehensive overview of the role of epigenetics in psychiatry, discussing its implications for genomics, evolution, inheritance patterns, and the development of biomarkers. Epigenetic mechanisms, including DNA methylation, histone modifications, and non-coding RNAs, are explored in the context of psychiatric conditions such as depression, schizophrenia, and post-traumatic stress disorder (PTSD). The discussion highlights how epigenetic modifications can influence brain dynamics, as measured by electroencephalography (EEG), and how artificial intelligence (AI) can accelerate the development of treatment options. The potential for epigenetic biomarkers to improve diagnosis, prognosis, and treatment is emphasized, along with the role of behavioral interventions in inducing beneficial epigenetic modifications. Future directions in epigenetic research, including the development of epigenetic therapies and the elucidation of molecular mechanisms underlying epigenetic inheritance, are also discussed. This article underscores the transformative potential of epigenetics in advancing our understanding of mental health and disease, paving the way for novel therapeutic interventions.

Clustered regularly interspaced short palindromic repeats (CRISPR) based gene-editing has begun to transform the treatment landscape of genetic diseases. The history of the discovery of CRISPR/CRISPR-associated (Cas) proteins/single guide RNA (sgRNA)-based gene-editing since the first report of repetitive sequences of unknown significance in 1987 is fascinating, instructive, and inspiring for future advances. The recent approval of CRISPR-Cas9-based gene therapy to treat patients with severe sickle cell anemia and transfusion-dependent beta thalassemia has renewed hope for treating other hematologic diseases, including patients with germline predisposition to hematologic malignancies, who would benefit greatly from the development of CRISPR-based gene therapies. The purpose of this manuscript is three-fold: first, a chronological description of the history of CRISPR-Cas9-sgRNA-based gene editing; second, a brief description of the current state of clinical research in hematologic diseases, including selected applications in treating hematologic diseases with CRISPR-based gene therapy; and third, the current progress in gene therapies in inherited hematologic diseases and bone marrow failure syndromes, to hopefully stimulate efforts towards developing these therapies for patients with inherited bone marrow failure syndromes and other inherited conditions with germline predisposition to hematologic malignancies.

The SARS (severe acute respiratory syndrome)-CoV (Coronavirus)-2 S(spike)-protein mRNA/cDNA currently being used as vaccines are antigenic but not antigens against SARS-CoV-2, that causes COVID (Coronavirus Disease) -19. Furthermore, the mRNA and cDNA antigenic vaccines also have potentials for homologous as well as heterologous recombination, primarily into the somatic cell DNA of the vaccine recipients. On the contrary, a SARS-CoV-2 RBD-protein antigen, a part of the S-protein, will directly stimulate antibody production against SARS-CoV-2. Hence, a vaccine composed of SARS-CoV-2 RBD-protein as a safer, fast acting, and effective vaccine against SARS-CoV-2 and thus against COVID-19. This is also useful for some immune compromised individuals.

Ultraviolet light incident on organic material can initiate its spontaneous dissipative structuring into chromophores which can catalyze their own replication. This may have been the case for one of the most ancient of all chromophores dissipating the Archean UVC photon flux, the nucleic acids. Oligos of nucleic acids with affinity to particular amino acids which foment UVC photon dissipation would have been selected through non-equilibrium thermodynamic imperatives which favor entropy production. Indeed, we show here that those amino acids with characteristics most relevant to fomenting UVC photon dissipation are precisely those with greatest chemical affinity to their codons or anticodons. Entropy production could thus provide an explanation for the accumulation of information in nucleic acids relevant to the dissipation of the externally imposed thermodynamic potentials. The accumulation of information in this manner provides a link between evolution and entropy production.

The limited information available on the structure of complexes involving transcription factors and cognate DNA response elements represents a major obstacle in the quest to understand their mechanism of action at the molecular level. We implemented a concerted structural proteomics approach, which combined hydrogen-deuterium exchange (HDX), quantitative protein-protein and protein-nucleic acid cross-linking (XL), and homology analysis, to model the structure of the complex between the full-length DNA binding domain (DBD) of FOXO4 and its DNA binding element (DBE). The results confirmed that FOXO4-DBD assumes the characteristic forkhead topology shared by these types of transcription factors, but its binding mode differs significantly from those of other members of the family. The results showed that the binding interaction stabilized regions that were rather flexible and disordered in the unbound form. Surprisingly, the conformational effects were not limited only to the interface between bound components but extended also to distal regions that may be essential to recruiting additional factors to the transcription machinery. In addition to providing valuable new insights into the binding mechanism, this project provided an excellent evaluation of the merits of structural proteomics approaches in the investigation of systems that arematerial not directly amenable to traditional high-resolution techniques.

Climate change has emerged as a crucial global issue that significantly threatens the survival of plants. In particular, low temperature (LT) is one of the critical environmental factors that influence plant morphological, physiological, and biochemical changes during both the vegetative and reproductive growth stages. LT, including abrupt drops in temperature, as well as winter conditions, can cause detrimental effects on the growth and development of tomato plants, ranging from sowing, transplanting, truss appearance, flowering, fertilization, flowering, fruit ripening, and yields. Therefore, it is imperative to understand the comprehensive mechanisms underlying the adaptation and acclimation of tomato plants to LT at the morphological and the molecular levels. In this review, we discuss the previous and current knowledge of morphological and physiological changes, which contains major factors and traits associated with the LT stress response during the period of the vegetative and reproductive growth and development in tomato plants. Moreover, we highlight recent advances in the understanding of physiological and biochemical responses, and molecular mechanisms, including LT perception, signaling transduction, gene regulation, and fruit ripening and epigenetic regulation. The comprehensive understanding of LT response provides a solid basis to develop the LT-resistant varieties for sustainable tomato production under the ever-changing temperature fluctuations.

Formulating a novel concept about the origin of human aging has been constrained by the dominance of a “classic theory” that was proposed nearly 70 years ago. Despite concern over the validity of some of its assumptions, the theory remained basic to our understanding of aging’s relationship with natural selection (NS). However, the logic upon which it rests was tested and subsequently challenged. The present theory describes the single cause of human aging consistent with Darwin’s evolutionary requirement for selection of adaptive traits. It describes an emergent property of the developmental program (DP), that is expressed upon completion of ontogenesis. It involves redundant expression of regulatory processes from the last stage of the DP. That mechanism subsequently preserves a non-aging, stable interval of unchanging NS during which reproductive fitness is achieved. Thereafter, loss of DP regulatory redundancy due to reliability limits, stochastic mutation accumulation, reproductive and a specific type of DNA damage, initiates aging which causes an inexorable decline in strength of NS to begin. It starts approximately a decade later than proposed in the classic theory. Since reproduction and aging are inextricably linked by the same emergent property, selection of that regulatory mechanism makes both traits products of NS.

The field of mitochondrial genomics has advanced rapidly and revolutionized disciplines from molecular anthropology, population genetics, to medical/oncogenetics. However, mtDNA next-generation sequencing (NGS) analysis for matrilineal haplotyping and phylogeographic inference remains hindered by the lack of a consolidated, mitogenome database and efficient bioinformatics pipeline. To address this, we developed a customized human mitogenome database (hMITO DB) embedded in a CLC Genomics workflow for read mapping, variant analysis, haplotyping, and geo-mapping. The database was constructed from 4,286 mitogenomes. The macro-haplogroup (A to Z) distribution and representative phylogenetic tree were found consistent with published literature. The hMITO DB automated workflow was tested using mtDNA-NGS sequences derived from Pap smears and cervical cancer cell lines. The auto-generated read mapping, variants track, and table of haplotypes and geo-origins were completed in 15 min for 47 samples. The mtDNA workflow proved to be a rapid, efficient and accurate means of sequence analysis for translational mitogenomics.

The World Health Organization identifies alcohol as a cause in several neoplasias of the oro-pharynx cavity, esophagus, gastrointestinal tract, larynx, liver, or female breast. This study re-views ethanol's nonoxidative and oxidative metabolism and one-carbon metabolism that en-compasses both redox and transfer reactions that influence crucial cell proliferation machinery. Ethanol favors the uncontrolled production and action of free radicals that interfere with the maintenance of essential cellular functions. We focus on the generation of protein, DNA, and lipid adducts that interfere with the cellular processes related to growth and differentiation. Ethanol's effects on stem cells responsible for building and repairing tissues are reviewed. Cancer stem cells suffer disturbances related to the expression of cell surface markers, enzymes, and transcription factors after ethanol exposure with consequent dysregulation of mechanisms related to cancer metastasis or resistance to treatments. Our analysis aims to underlie and discuss potential targets that show more sensitivity to ethanol's action and identify specific metabolic routes and metabolic realms that may be corrected to recover metabolic homeostasis after pharmacological interven-tion. Specifically, research should pay attention to reestablish metabolic fluxes by fine-tuning the functioning of specific pathways related to one-carbon metabolism and antioxidant processes.

The Third U.S. National Marine Environmental DNA Workshop on June 2–5, 2024 brought together researchers, practitioners, and policymakers to overview and discuss: (1) environmental (e)DNA – defined as “DNA in the environment” – as a strategic U.S. national priority, (2) eDNA tool readiness and approaches to support decision-making, (3) emerging eDNA technologies, and (4) plans for implementation and adoption for environmental monitoring and ecosystem assessment “from microbes through whales”. Uses and applications of eDNA have been revolutionizing exploration, measurement, and monitoring of biodiversity across ecosystems, including the marine and freshwater aquatic environments emphasized in the Workshop, as well as terrestrial and aerial systems. The Workshop featured the launch of the new U.S. National Aquatic Environmental DNA Strategy from the White House Office of Science and Technology Policy (2024), with aquatic defined as freshwater to ocean waters, across all salinities; the Strategy was developed using outputs and a taskforce from the 2022 Second Workshop. The 2.5-day Third Workshop included speaker presentations, panel discussions, and networking on eDNA tools, applications, use, upscaling, and their future. The Workshop led into Capitol Hill Ocean Week (CHOW) events (June 5–6, 2024), during World Ocean Week. Note that this summary paper represents the author’s take-away view and interpretation, and not necessarily the overall views or consensus of the U.S. Government, Smithsonian Institution, workshop organizers, or speakers.

Finding out how cells with the same genome change fates in differentiation commitment is a challenge of biology. We used MCF-7 breast cancer cells treated with the ErbB2 ligand heregulin (HRG), which induces differentiation, to address if and how the constitutive pericentromere-associated domains (PADs) may be involved in this process. PAD-specific repressive heterochromatin (H3K9me3) and active euchromatin (H3K4me3) marking, centromere (CENPA) labelling, qPCR, acridine-orange-DNA structural test, and microscopic image analysis were applied. We found a two-step DNA unfolding, at 15-20 min and 60 min after HRG treatment, coinciding with bi-phasic activation of the early response genes (c-FOS family) and two steps of critical phase transition which were revealed in transcriptome studies. In control, the distribution of PAD number and size displays a power-law scaling with a boundary at the nucleolus. PADs’ clustering correlates with centromere numbers. 15 min after HRG treatment, the unravelling of PADs occurs, coinciding with the first step of euchromatin unfolding. The second step is associated with transcription of long-non-coding-RNA from satellite III DNA. We hypothesize that splitting of the PAD clusters under the critical size threshold of the silencing domain abrupts position effect variegation. It allows the first genome transcription avalanche to occur, starting differentiation commitment.