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Spatial analysis of Cdc42 activity reveals a role for plasma membrane–associated Cdc42 in centrosome regulation

The ability of the small GTPase Cdc42 to regulate diverse cellular processes depends on tight spatial control of its activity. Cdc42 function is best understood at the plasma membrane (PM), where it regulates cytoskeletal organization and cell polarization. Active Cdc42 has also been detected at the...

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Autores principales: Herrington, Kari A., Trinh, Andrew L., Dang, Carolyn, O’Shaughnessy, Ellen, Hahn, Klaus M., Gratton, Enrico, Digman, Michelle A., Sütterlin, Christine
Formato: Online Artículo Texto
Lenguaje:English
Publicado: The American Society for Cell Biology 2017
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5509425/
https://www.ncbi.nlm.nih.gov/pubmed/28539409
http://dx.doi.org/10.1091/mbc.E16-09-0665
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author Herrington, Kari A.
Trinh, Andrew L.
Dang, Carolyn
O’Shaughnessy, Ellen
Hahn, Klaus M.
Gratton, Enrico
Digman, Michelle A.
Sütterlin, Christine
author_facet Herrington, Kari A.
Trinh, Andrew L.
Dang, Carolyn
O’Shaughnessy, Ellen
Hahn, Klaus M.
Gratton, Enrico
Digman, Michelle A.
Sütterlin, Christine
author_sort Herrington, Kari A.
collection PubMed
description The ability of the small GTPase Cdc42 to regulate diverse cellular processes depends on tight spatial control of its activity. Cdc42 function is best understood at the plasma membrane (PM), where it regulates cytoskeletal organization and cell polarization. Active Cdc42 has also been detected at the Golgi, but its role and regulation at this organelle are only partially understood. Here we analyze the spatial distribution of Cdc42 activity by moni­toring the dynamics of the Cdc42 FLARE biosensor using the phasor approach to FLIM-FRET. Phasor analysis revealed that Cdc42 is active at all Golgi cisternae and that this activity is controlled by Tuba and ARHGAP10, two Golgi-associated Cdc42 regulators. To our surprise, FGD1, another Cdc42 GEF at the Golgi, was not required for Cdc42 regulation at the Golgi, although its depletion decreased Cdc42 activity at the PM. Similarly, changes in Golgi morphology did not affect Cdc42 activity at the Golgi but were associated with a substantial reduction in PM-associated Cdc42 activity. Of interest, cells with reduced Cdc42 activity at the PM displayed altered centrosome morphology, suggesting that centrosome regulation may be mediated by active Cdc42 at the PM. Our study describes a novel quantitative approach to determine Cdc42 activity at specific subcellular locations and reveals new regulatory principles and functions of this small GTPase.
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spelling pubmed-55094252017-09-30 Spatial analysis of Cdc42 activity reveals a role for plasma membrane–associated Cdc42 in centrosome regulation Herrington, Kari A. Trinh, Andrew L. Dang, Carolyn O’Shaughnessy, Ellen Hahn, Klaus M. Gratton, Enrico Digman, Michelle A. Sütterlin, Christine Mol Biol Cell Articles The ability of the small GTPase Cdc42 to regulate diverse cellular processes depends on tight spatial control of its activity. Cdc42 function is best understood at the plasma membrane (PM), where it regulates cytoskeletal organization and cell polarization. Active Cdc42 has also been detected at the Golgi, but its role and regulation at this organelle are only partially understood. Here we analyze the spatial distribution of Cdc42 activity by moni­toring the dynamics of the Cdc42 FLARE biosensor using the phasor approach to FLIM-FRET. Phasor analysis revealed that Cdc42 is active at all Golgi cisternae and that this activity is controlled by Tuba and ARHGAP10, two Golgi-associated Cdc42 regulators. To our surprise, FGD1, another Cdc42 GEF at the Golgi, was not required for Cdc42 regulation at the Golgi, although its depletion decreased Cdc42 activity at the PM. Similarly, changes in Golgi morphology did not affect Cdc42 activity at the Golgi but were associated with a substantial reduction in PM-associated Cdc42 activity. Of interest, cells with reduced Cdc42 activity at the PM displayed altered centrosome morphology, suggesting that centrosome regulation may be mediated by active Cdc42 at the PM. Our study describes a novel quantitative approach to determine Cdc42 activity at specific subcellular locations and reveals new regulatory principles and functions of this small GTPase. The American Society for Cell Biology 2017-07-15 /pmc/articles/PMC5509425/ /pubmed/28539409 http://dx.doi.org/10.1091/mbc.E16-09-0665 Text en © 2017 Herrington et al. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0). “ASCB®,” “The American Society for Cell Biology®,” and “Molecular Biology of the Cell®” are registered trademarks of The American Society for Cell Biology.
spellingShingle Articles
Herrington, Kari A.
Trinh, Andrew L.
Dang, Carolyn
O’Shaughnessy, Ellen
Hahn, Klaus M.
Gratton, Enrico
Digman, Michelle A.
Sütterlin, Christine
Spatial analysis of Cdc42 activity reveals a role for plasma membrane–associated Cdc42 in centrosome regulation
title Spatial analysis of Cdc42 activity reveals a role for plasma membrane–associated Cdc42 in centrosome regulation
title_full Spatial analysis of Cdc42 activity reveals a role for plasma membrane–associated Cdc42 in centrosome regulation
title_fullStr Spatial analysis of Cdc42 activity reveals a role for plasma membrane–associated Cdc42 in centrosome regulation
title_full_unstemmed Spatial analysis of Cdc42 activity reveals a role for plasma membrane–associated Cdc42 in centrosome regulation
title_short Spatial analysis of Cdc42 activity reveals a role for plasma membrane–associated Cdc42 in centrosome regulation
title_sort spatial analysis of cdc42 activity reveals a role for plasma membrane–associated cdc42 in centrosome regulation
topic Articles
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5509425/
https://www.ncbi.nlm.nih.gov/pubmed/28539409
http://dx.doi.org/10.1091/mbc.E16-09-0665
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