The interplay of Rac1 activity, ubiquitination and GDI binding and its consequences for endothelial cell spreading.

Signaling by the Rho GTPase Rac1 is key to the regulation of cytoskeletal dynamics, cell spreading and adhesion. It is widely accepted that the inactive form of Rac1 is bound by Rho GDI, which prevents Rac1 activation and Rac1-effector interactions. In addition, GDI-bound Rac1 is protected from prot...

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Autores principales:	Jisca Majolée, Fabienne Podieh, Peter L Hordijk, Igor Kovačević
Formato:	article
Lenguaje:	EN
Publicado:	Public Library of Science (PLoS) 2021
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/d63de637915a493c937ec2973bb473a3
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spelling	oai:doaj.org-article:d63de637915a493c937ec2973bb473a32021-12-02T20:09:16ZThe interplay of Rac1 activity, ubiquitination and GDI binding and its consequences for endothelial cell spreading.1932-620310.1371/journal.pone.0254386https://doaj.org/article/d63de637915a493c937ec2973bb473a32021-01-01T00:00:00Zhttps://doi.org/10.1371/journal.pone.0254386https://doaj.org/toc/1932-6203Signaling by the Rho GTPase Rac1 is key to the regulation of cytoskeletal dynamics, cell spreading and adhesion. It is widely accepted that the inactive form of Rac1 is bound by Rho GDI, which prevents Rac1 activation and Rac1-effector interactions. In addition, GDI-bound Rac1 is protected from proteasomal degradation, in line with data showing that Rac1 ubiquitination occurs exclusively when Rac1 is activated. We set out to investigate how Rac1 activity, GDI binding and ubiquitination are linked. We introduced single amino acid mutations in Rac1 which differentially altered Rac1 activity, and compared whether the level of Rac1 activity relates to Rac1 ubiquitination and GDI binding. Results show that Rac1 ubiquitination and the active Rac1 morphology is proportionally increased with Rac1 activity. Similarly, we introduced lysine-to-arginine mutations in constitutively active Rac1 to inhibit site-specific ubiquitination and analyze this effect on Rac1 signaling output and ubiquitination. These data show that the K16R mutation inhibits GTP binding, and consequently Rac1 activation, signaling and-ubiquitination, while the K147R mutation does not block Rac1 signaling, but does inhibits its ubiquitination. In both sets of mutants, no direct correlation was observed between GDI binding and Rac1 activity or -ubiquitination. Taken together, our data show that a strong, positive correlation exists between Rac1 activity and its level of ubiquitination, but also that GDI dissociation does not predispose Rac1 to ubiquitination.Jisca MajoléeFabienne PodiehPeter L HordijkIgor KovačevićPublic Library of Science (PLoS)articleMedicineRScienceQENPLoS ONE, Vol 16, Iss 7, p e0254386 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Jisca Majolée Fabienne Podieh Peter L Hordijk Igor Kovačević The interplay of Rac1 activity, ubiquitination and GDI binding and its consequences for endothelial cell spreading.
description	Signaling by the Rho GTPase Rac1 is key to the regulation of cytoskeletal dynamics, cell spreading and adhesion. It is widely accepted that the inactive form of Rac1 is bound by Rho GDI, which prevents Rac1 activation and Rac1-effector interactions. In addition, GDI-bound Rac1 is protected from proteasomal degradation, in line with data showing that Rac1 ubiquitination occurs exclusively when Rac1 is activated. We set out to investigate how Rac1 activity, GDI binding and ubiquitination are linked. We introduced single amino acid mutations in Rac1 which differentially altered Rac1 activity, and compared whether the level of Rac1 activity relates to Rac1 ubiquitination and GDI binding. Results show that Rac1 ubiquitination and the active Rac1 morphology is proportionally increased with Rac1 activity. Similarly, we introduced lysine-to-arginine mutations in constitutively active Rac1 to inhibit site-specific ubiquitination and analyze this effect on Rac1 signaling output and ubiquitination. These data show that the K16R mutation inhibits GTP binding, and consequently Rac1 activation, signaling and-ubiquitination, while the K147R mutation does not block Rac1 signaling, but does inhibits its ubiquitination. In both sets of mutants, no direct correlation was observed between GDI binding and Rac1 activity or -ubiquitination. Taken together, our data show that a strong, positive correlation exists between Rac1 activity and its level of ubiquitination, but also that GDI dissociation does not predispose Rac1 to ubiquitination.
format	article
author	Jisca Majolée Fabienne Podieh Peter L Hordijk Igor Kovačević
author_facet	Jisca Majolée Fabienne Podieh Peter L Hordijk Igor Kovačević
author_sort	Jisca Majolée
title	The interplay of Rac1 activity, ubiquitination and GDI binding and its consequences for endothelial cell spreading.
title_short	The interplay of Rac1 activity, ubiquitination and GDI binding and its consequences for endothelial cell spreading.
title_full	The interplay of Rac1 activity, ubiquitination and GDI binding and its consequences for endothelial cell spreading.
title_fullStr	The interplay of Rac1 activity, ubiquitination and GDI binding and its consequences for endothelial cell spreading.
title_full_unstemmed	The interplay of Rac1 activity, ubiquitination and GDI binding and its consequences for endothelial cell spreading.
title_sort	interplay of rac1 activity, ubiquitination and gdi binding and its consequences for endothelial cell spreading.
publisher	Public Library of Science (PLoS)
publishDate	2021
url	https://doaj.org/article/d63de637915a493c937ec2973bb473a3
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The interplay of Rac1 activity, ubiquitination and GDI binding and its consequences for endothelial cell spreading.

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