title: The guanine-nucleotide-exchange factor P-Rex1 is activated by protein phosphatase 1α.
abstract: 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.
authors.affiliation: Inositide Programme, Babraham Institute, Cambridge, UK.
authors.forename: Mark A
authors.initials: MA
authors.lastname: Barber
authors.forename: Annick
authors.initials: A
authors.lastname: Hendrickx
authors.forename: Monique
authors.initials: M
authors.lastname: Beullens
authors.forename: Hugo
authors.initials: H
authors.lastname: Ceulemans
authors.forename: David
authors.initials: D
authors.lastname: Oxley
authors.forename: Sylvia
authors.initials: S
authors.lastname: Thelen
authors.forename: Marcus
authors.initials: M
authors.lastname: Thelen
authors.forename: Mathieu
authors.initials: M
authors.lastname: Bollen
authors.forename: Heidi C E
authors.initials: HC
authors.lastname: Welch
mesh_terms: D020816:Amino Acid Motifs; D000818:Animals; D001011:Aorta / Q000166:cytology; D048430:Cell Shape; D002478:Cells, Cultured; D042783:Endothelial Cells / Q000187:drug effects / Q000378:metabolism / Q000502:physiology; D020662:Guanine Nucleotide Exchange Factors / Q000737:chemistry*; D006801:Humans; D010766:Phosphorylation; D010982:Platelet-Derived Growth Factor / Q000494:pharmacology / Q000502:physiology; D011485:Protein Binding; D054645:Protein Phosphatase 1 / Q000737:chemistry* / Q000378:metabolism; D017434:Protein Structure, Tertiary; D011817:Rabbits; D013552:Swine; D020830:rac1 GTP-Binding Protein / Q000378:metabolism
chemical_list: D020662:Guanine Nucleotide Exchange Factors; C453280:PREX1 protein, human; C490712:PREX2 protein, human; D010982:Platelet-Derived Growth Factor; C107953:RAC1 protein, human; D054645:Protein Phosphatase 1; D020830:rac1 GTP-Binding Protein