Secretory pathway-dependent localization of the Saccharomyces cerevisiae Rho GTPase-activating protein Rgd1p at growth sites

Eukaryot Cell. 2012 May;11(5):590-600. doi: 10.1128/EC.00042-12. Epub 2012 Mar 23.

Abstract

Establishment and maintenance of cell polarity in eukaryotes depends upon the regulation of Rho GTPases. In Saccharomyces cerevisiae, the Rho GTPase activating protein (RhoGAP) Rgd1p stimulates the GTPase activities of Rho3p and Rho4p, which are involved in bud growth and cytokinesis, respectively. Consistent with the distribution of Rho3p and Rho4p, Rgd1p is found mostly in areas of polarized growth during cell cycle progression. Rgd1p was mislocalized in mutants specifically altered for Golgi apparatus-based phosphatidylinositol 4-P [PtdIns(4)P] synthesis and for PtdIns(4,5)P(2) production at the plasma membrane. Analysis of Rgd1p distribution in different membrane-trafficking mutants suggested that Rgd1p was delivered to growth sites via the secretory pathway. Rgd1p may associate with post-Golgi vesicles by binding to PtdIns(4)P and then be transported by secretory vesicles to the plasma membrane. In agreement, we show that Rgd1p coimmunoprecipitated and localized with markers specific to secretory vesicles and cofractionated with a plasma membrane marker. Moreover, in vivo imaging revealed that Rgd1p was transported in an anterograde manner from the mother cell to the daughter cell in a vectoral manner. Our data indicate that secretory vesicles are involved in the delivery of RhoGAP Rgd1p to the bud tip and bud neck.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Cell Membrane / metabolism
  • Cell Polarity
  • Electrophoresis, Polyacrylamide Gel
  • GTPase-Activating Proteins / metabolism*
  • Golgi Apparatus / metabolism
  • Green Fluorescent Proteins / metabolism
  • Guanine Nucleotide Exchange Factors / metabolism
  • Immunoprecipitation
  • Phosphatidylinositol 4,5-Diphosphate / biosynthesis
  • Phosphatidylinositol 4,5-Diphosphate / genetics
  • Phosphatidylinositol Phosphates / biosynthesis
  • Phosphatidylinositol Phosphates / genetics
  • Plasmids / metabolism
  • Protein Binding
  • Protein Transport
  • Saccharomyces cerevisiae / enzymology
  • Saccharomyces cerevisiae / genetics
  • Saccharomyces cerevisiae / growth & development
  • Saccharomyces cerevisiae / metabolism*
  • Saccharomyces cerevisiae Proteins / metabolism*
  • Secretory Pathway*
  • Secretory Vesicles / metabolism*
  • rab GTP-Binding Proteins / metabolism

Substances

  • GTPase-Activating Proteins
  • Guanine Nucleotide Exchange Factors
  • Phosphatidylinositol 4,5-Diphosphate
  • Phosphatidylinositol Phosphates
  • RGD1 protein, S cerevisiae
  • SEC2 protein, S cerevisiae
  • Saccharomyces cerevisiae Proteins
  • phosphatidylinositol 4-phosphate
  • Green Fluorescent Proteins
  • SEC4 protein, S cerevisiae
  • rab GTP-Binding Proteins