title: Protein kinase C-theta (PKCθ) phosphorylates and inhibits the guanine exchange factor, GIV/Girdin.
abstract: Gα-interacting, vesicle-associated protein (GIV/Girdin) is a multidomain signal transducer that enhances PI3K-Akt signals downstream of both G-protein-coupled receptors and growth factor receptor tyrosine kinases during diverse biological processes and cancer metastasis. Mechanistically, GIV serves as a non-receptor guanine nucleotide exchange factor (GEF) that enhances PI3K signals by activating trimeric G proteins, Gαi1/2/3. Site-directed mutations in GIV's GEF motif disrupt its ability to bind or activate Gi and abrogate PI3K-Akt signals; however, nothing is known about how GIV's GEF function is regulated. Here we report that PKCθ, a novel protein kinase C, down-regulates GIV's GEF function by phosphorylating Ser(S)1689 located within GIV's GEF motif. We demonstrate that PKCθ specifically binds and phosphorylates GIV at S1689, and this phosphoevent abolishes GIV's ability to bind and activate Gαi. HeLa cells stably expressing the phosphomimetic mutant of GIV, GIV-S1689→D, are phenotypically identical to those expressing the GEF-deficient F1685A mutant: Actin stress fibers are decreased and cell migration is inhibited whereas cell proliferation is triggered, and Akt (a.k.a. protein kinase B, PKB) activation is impaired downstream of both the lysophosphatidic acid receptor, a G-protein-coupled receptor, and the insulin receptor, a receptor tyrosine kinase. These findings indicate that phosphorylation of GIV by PKCθ inhibits GIV's GEF function and generates a unique negative feedback loop for downregulating the GIV-Gi axis of prometastatic signaling downstream of multiple ligand-activated receptors. This phosphoevent constitutes the only regulatory pathway described for terminating signaling by any of the growing family of nonreceptor GEFs that modulate G-protein activity.
authors.affiliation: Department of Medicine, University of California at San Diego School of Medicine, La Jolla, CA 92093, USA.
authors.forename: Inmaculada
authors.initials: I
authors.lastname: López-Sánchez
authors.forename: Mikel
authors.initials: M
authors.lastname: Garcia-Marcos
authors.forename: Yash
authors.initials: Y
authors.lastname: Mittal
authors.forename: Nicolas
authors.initials: N
authors.lastname: Aznar
authors.forename: Marilyn G
authors.initials: MG
authors.lastname: Farquhar
authors.forename: Pradipta
authors.initials: P
authors.lastname: Ghosh
mesh_terms: D000199:Actins / Q000378:metabolism; D020662:Guanine Nucleotide Exchange Factors / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D015151:Immunoblotting; D047468:Immunoprecipitation; D007527:Isoenzymes / Q000378:metabolism*; D008840:Microfilament Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D008958:Models, Molecular; D020125:Mutation, Missense / Q000235:genetics; D010766:Phosphorylation; D011493:Protein Kinase C / Q000378:metabolism*; D000074121:Protein Kinase C-theta; D015398:Signal Transduction / Q000235:genetics*; D033921:Vesicular Transport Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*
chemical_list: D000199:Actins; C502451:CCDC88A protein, human; D020662:Guanine Nucleotide Exchange Factors; D007527:Isoenzymes; D008840:Microfilament Proteins; D033921:Vesicular Transport Proteins; C076233:PRKCQ protein, human; D011493:Protein Kinase C; D000074121:Protein Kinase C-theta