P-Rex1 is a GEF (guanine-nucleotide-exchange factor) for the small G-protein Rac that is activated by PIP3 (phosphatidylinositol 3,4,5-trisphosphate) and Gβγ subunits and inhibited by PKA (protein kinase A). In the present study we show that PP1α (protein phosphatase 1α) binds P-Rex1 through an RVxF-type docking motif. PP1α activates P-Rex1 directly in vitro, both independently of and additively to PIP3 and Gβγ. PP1α also substantially activates P-Rex1 in vivo, both in basal and PDGF (platelet-derived growth factor)- or LPA (lysophosphatidic acid)-stimulated cells. The phosphatase activity of PP1α is required for P-Rex1 activation. PP1β, a close homologue of PP1α, is also able to activate P-Rex1, but less effectively. PP1α stimulates P-Rex1-mediated Rac-dependent changes in endothelial cell morphology. MS analysis of wild-type P-Rex1 and a PP1α-binding-deficient mutant revealed that endogenous PP1α dephosphorylates P-Rex1 on at least three residues, Ser834, Ser1001 and Ser1165. Site-directed mutagenesis of Ser1165 to alanine caused activation of P-Rex1 to a similar degree as did PP1α, confirming Ser1165 as a dephosphorylation site important in regulating P-Rex1 Rac-GEF activity. In summary, we have identified a novel mechanism for direct activation of P-Rex1 through PP1α-dependent dephosphorylation.
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Research Article|
March 14 2012
The guanine-nucleotide-exchange factor P-Rex1 is activated by protein phosphatase 1α
Mark A. Barber;
Mark A. Barber
*Inositide Programme, Babraham Institute, Cambridge, U.K.
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Annick Hendrickx;
Annick Hendrickx
†Department of Molecular Cell Biology, Catholic University Leuven, Leuven, Belgium
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Monique Beullens;
Monique Beullens
†Department of Molecular Cell Biology, Catholic University Leuven, Leuven, Belgium
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Hugo Ceulemans;
Hugo Ceulemans
†Department of Molecular Cell Biology, Catholic University Leuven, Leuven, Belgium
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David Oxley;
David Oxley
‡Proteomics Group, Babraham Institute, Cambridge, U.K.
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Sylvia Thelen;
Sylvia Thelen
§Institute for Research in Biomedicine, Bellinzona, Switzerland
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Marcus Thelen;
Marcus Thelen
§Institute for Research in Biomedicine, Bellinzona, Switzerland
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Mathieu Bollen;
Mathieu Bollen
†Department of Molecular Cell Biology, Catholic University Leuven, Leuven, Belgium
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Heidi C. E. Welch
Heidi C. E. Welch
1
*Inositide Programme, Babraham Institute, Cambridge, U.K.
1To whom correspondence should be addressed (email heidi.welch@babraham.ac.uk).
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Publisher: Portland Press Ltd
Received:
November 28 2011
Revision Received:
January 10 2012
Accepted:
January 16 2012
Accepted Manuscript online:
January 16 2012
Online ISSN: 1470-8728
Print ISSN: 0264-6021
© The Authors Journal compilation © 2012 Biochemical Society
2012
Biochem J (2012) 443 (1): 173–183.
Article history
Received:
November 28 2011
Revision Received:
January 10 2012
Accepted:
January 16 2012
Accepted Manuscript online:
January 16 2012
Citation
Mark A. Barber, Annick Hendrickx, Monique Beullens, Hugo Ceulemans, David Oxley, Sylvia Thelen, Marcus Thelen, Mathieu Bollen, Heidi C. E. Welch; The guanine-nucleotide-exchange factor P-Rex1 is activated by protein phosphatase 1α. Biochem J 1 April 2012; 443 (1): 173–183. doi: https://doi.org/10.1042/BJ20112078
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