Version 1
: Received: 27 August 2024 / Approved: 27 August 2024 / Online: 28 August 2024 (09:38:56 CEST)
How to cite:
Shin, Y.-C.; Cho, M.; Hwang, J. M.; Myung, K.; Kweon, H.-S.; Seong, H.-A.; Lee, Z.-W.; Lee, K.-B. Imaging the Raf–MEK–ERK Signaling Cascade in Living Cells. Preprints2024, 2024082028. https://doi.org/10.20944/preprints202408.2028.v1
Shin, Y.-C.; Cho, M.; Hwang, J. M.; Myung, K.; Kweon, H.-S.; Seong, H.-A.; Lee, Z.-W.; Lee, K.-B. Imaging the Raf–MEK–ERK Signaling Cascade in Living Cells. Preprints 2024, 2024082028. https://doi.org/10.20944/preprints202408.2028.v1
Shin, Y.-C.; Cho, M.; Hwang, J. M.; Myung, K.; Kweon, H.-S.; Seong, H.-A.; Lee, Z.-W.; Lee, K.-B. Imaging the Raf–MEK–ERK Signaling Cascade in Living Cells. Preprints2024, 2024082028. https://doi.org/10.20944/preprints202408.2028.v1
APA Style
Shin, Y. C., Cho, M., Hwang, J. M., Myung, K., Kweon, H. S., Seong, H. A., Lee, Z. W., & Lee, K. B. (2024). Imaging the Raf–MEK–ERK Signaling Cascade in Living Cells. Preprints. https://doi.org/10.20944/preprints202408.2028.v1
Chicago/Turabian Style
Shin, Y., Zee-Won Lee and Kyung-Bok Lee. 2024 "Imaging the Raf–MEK–ERK Signaling Cascade in Living Cells" Preprints. https://doi.org/10.20944/preprints202408.2028.v1
Abstract
Conventional biochemical methods for studying cellular signaling cascades have relied on destructive cell disruption. In contrast, live cell imaging of fluorescent-tagged transfected proteins offers a non-invasive approach to understanding signal transduction events. One strategy involves monitoring the phosphorylation-dependent shuttling of a fluorescent-labeled kinase between the nucleus and cytoplasm using nuclear localization, export signals, or both. This paper introduces a simple method to visualize intracellular signal transduction in live cells by exploring the translocation properties of PKC from the cytoplasm to the membrane. We fused bait protein to PKC, allowing the bait (RFP-labeled) and target (GFP-labeled) proteins to co-translocate from the cytoplasm to the membrane. However, in non-interacting protein pairs, only the bait protein was translocated to the plasma membrane. To verify our approach, we examined the Raf–MEK–ERK signaling cascade (ERK pathway) and directly observed the Raf1/MEK2 interaction. We also identified ternary complexes containing the KSR1 scaffold protein. Additionally, the MEK/ERK interaction was dependent on the exogenous expression of KSR1.
Biology and Life Sciences, Biology and Biotechnology
Copyright:
This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
