title: Phosphorylation of the Rab exchange factor Sec2p directs a switch in regulatory binding partners.
abstract: Sec2p is a guanine nucleotide exchange factor that promotes exocytosis by activating the Rab GTPase Sec4p. Sec2p is highly phosphorylated, and we have explored the role of phosphorylation in the regulation of its function. We have identified three phosphosites and demonstrate that phosphorylation regulates the interaction of Sec2p with its binding partners Ypt32p, Sec15p, and phosphatidyl-inositol-4-phosphate. In its nonphosphorylated form, Sec2p binds preferentially to the upstream Rab, Ypt32p-GTP, thus forming a Rab guanine nucleotide exchange factor cascade that leads to the activation of the downstream Rab, Sec4p. The nonphosphorylated form of Sec2p also binds to the Golgi-associated phosphatidyl-inositol-4-phosphate, which works in concert with Ypt32p-GTP to recruit Sec2p to Golgi-derived secretory vesicles. In contrast, the phosphorylated form of Sec2p binds preferentially to Sec15p, a downstream effector of Sec4p and a component of the exocyst tethering complex, thus forming a positive-feedback loop that prepares the secretory vesicle for fusion with the plasma membrane. Our results suggest that the phosphorylation state of Sec2p can direct a switch in its regulatory binding partners that facilitates maturation of the secretory vesicle and helps to promote the directionality of vesicular transport.
authors.affiliation: Department of Cellular and Molecular Medicine and Department of Chemistry and Biochemistry, Biomolecular and Proteomics Mass Spectrometry Facility, University of California, San Diego, La Jolla, CA 92093.
authors.forename: Danièle
authors.initials: D
authors.lastname: Stalder
authors.forename: Emi
authors.initials: E
authors.lastname: Mizuno-Yamasaki
authors.forename: Majid
authors.initials: M
authors.lastname: Ghassemian
authors.forename: Peter J
authors.initials: PJ
authors.lastname: Novick
mesh_terms: D001692:Biological Transport / Q000502:physiology; D004591:Electrophoresis, Polyacrylamide Gel; D020662:Guanine Nucleotide Exchange Factors / Q000235:genetics / Q000378:metabolism*; D047468:Immunoprecipitation; D008854:Microscopy, Electron; D008856:Microscopy, Fluorescence; D016297:Mutagenesis, Site-Directed; D018129:Phosphatidylinositol Phosphates / Q000378:metabolism; D010766:Phosphorylation; D011485:Protein Binding; D012441:Saccharomyces cerevisiae / Q000378:metabolism / Q000502:physiology*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*; D022161:Transport Vesicles / Q000378:metabolism*; D033921:Vesicular Transport Proteins / Q000378:metabolism; D020691:rab GTP-Binding Proteins / Q000378:metabolism*
chemical_list: D020662:Guanine Nucleotide Exchange Factors; D018129:Phosphatidylinositol Phosphates; C060862:SEC15 protein, S cerevisiae; C064456:SEC2 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D033921:Vesicular Transport Proteins; C037178:phosphatidylinositol 4-phosphate; C121633:YPT32 protein, S cerevisiae; D020691:rab GTP-Binding Proteins
keywords: membrane traffic; phospho-regulation; vesicle maturation; yeast