Spatiotemporal dynamics of GEF-H1 activation controlled by microtubule- and Src-mediated pathways
| Autor: | Daniel J. Marston, Philippe Roudot, Mitsu Ikura, Timothy A. Daugird, Christopher B. Marshall, Robert Rottapel, Jungsik Noh, María José Sandí, Sidney L. Lisanza, Klaus M. Hahn, John Sondek, Gaudenz Danuser, Mihai L. Azoitei |
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| Jazyk: | angličtina |
| Rok vydání: | 2019 |
| Předmět: |
animal structures
RHOA GTPase macromolecular substances Biosensing Techniques environment and public health Microtubules Article 03 medical and health sciences 0302 clinical medicine Microtubule Cell cortex Chlorocebus aethiops Animals Humans Cytoskeleton Research Articles 030304 developmental biology 0303 health sciences biology fungi technology industry and agriculture Cell Biology enzymes and coenzymes (carbohydrates) HEK293 Cells src-Family Kinases COS Cells Biophysics biology.protein Guanine nucleotide exchange factor biological phenomena cell phenomena and immunity Signal transduction rhoA GTP-Binding Protein 030217 neurology & neurosurgery Rho Guanine Nucleotide Exchange Factors Proto-oncogene tyrosine-protein kinase Src Signal Transduction |
| Zdroj: | The Journal of Cell Biology |
| ISSN: | 1540-8140 0021-9525 |
| Popis: | Azoitei, Noh, et al. engineer fluorescent biosensors to measure activation at the subcellular level and with subsecond kinetics of GEF-H1, a Rho GTPase that regulates cytoskeletal dynamics. In combination with computational image time series analysis, the biosensors reveal the synergistic role of microtubule dynamics and Src phosphorylation in regulating GEF-H1 activity locally during cell migration. Rho family GTPases are activated with precise spatiotemporal control by guanine nucleotide exchange factors (GEFs). Guanine exchange factor H1 (GEF-H1), a RhoA activator, is thought to act as an integrator of microtubule (MT) and actin dynamics in diverse cell functions. Here we identify a GEF-H1 autoinhibitory sequence and exploit it to produce an activation biosensor to quantitatively probe the relationship between GEF-H1 conformational change, RhoA activity, and edge motion in migrating cells with micrometer- and second-scale resolution. Simultaneous imaging of MT dynamics and GEF-H1 activity revealed that autoinhibited GEF-H1 is localized to MTs, while MT depolymerization subadjacent to the cell cortex promotes GEF-H1 activation in an ~5-µm-wide peripheral band. GEF-H1 is further regulated by Src phosphorylation, activating GEF-H1 in a narrower band ~0–2 µm from the cell edge, in coordination with cell protrusions. This indicates a synergistic intersection between MT dynamics and Src signaling in RhoA activation through GEF-H1. |
| Databáze: | OpenAIRE |
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