Spatial regulation of the exocyst complex by Rho1 GTPase

Spatial regulation of membrane traffic is fundamental to many biological processes, including epithelial cell polarization and neuronal synaptogenesis. The multiprotein exocyst complex is localized to sites of polarized exocytosis, and is required for vesicle targeting and docking at specific domains of the plasma membrane. One component of the complex, Sec3, is thought to be a spatial landmark for polarized exocytosis. We have searched for proteins that regulate the polarized localization of the exocyst in the budding yeast Saccharomyces cerevisiae. Here we report that certain rho1 mutant alleles specifically affect the localization of the exocyst proteins. Sec3 interacts directly with Rho1 in its GTP-bound form, and functional Rho1 is needed both to establish and to maintain the polarized localization of Sec3. Sec3 is not the only mediator of the effect of Rho1 on the exocyst, because some members of the complex are correctly targeted independently of the interaction between Rho1 and Sec3. These results reveal the action of parallel pathways for the polarized localization of the exocytic machinery, both of which are under the control of Rho1, a master regulator of cell polarity.

[1]  P. Novick,et al.  Spatial Regulation of Exocytosis: Lessons from Yeast , 1998, The Journal of cell biology.

[2]  E. Cabib,et al.  The Gtp-Binding Protein Rho1p Is Required for Cell Cycle Progression and Polarization of the Yeast Cell , 1999, The Journal of cell biology.

[3]  P. Novick,et al.  The Exocyst is a multiprotein complex required for exocytosis in Saccharomyces cerevisiae. , 1996, The EMBO journal.

[4]  P. Novick,et al.  Sec6, Sec8, and Sec15 are components of a multisubunit complex which localizes to small bud tips in Saccharomyces cerevisiae , 1995, The Journal of cell biology.

[5]  Joan E. Adamo,et al.  The Rho GTPase Rho3 has a direct role in exocytosis that is distinct from its role in actin polarity. , 1999, Molecular biology of the cell.

[6]  Navin Pokala,et al.  High Rates of Actin Filament Turnover in Budding Yeast and Roles for Actin in Establishment and Maintenance of Cell Polarity Revealed Using the Actin Inhibitor Latrunculin-A , 1997, The Journal of cell biology.

[7]  P. Novick,et al.  Exo84p Is an Exocyst Protein Essential for Secretion* , 1999, The Journal of Biological Chemistry.

[8]  M. Götte,et al.  Vesicular transport: how many Ypt/Rab-GTPases make a eukaryotic cell? , 1997, Trends in biochemical sciences.

[9]  David Botstein,et al.  Phenotypic Analysis of Temperature-sensitive Yeast Actin Mutants , 2022 .

[10]  K. Tanaka,et al.  Growth site localization of Rho1 small GTP-binding protein and its involvement in bud formation in Saccharomyces cerevisiae , 1994, The Journal of cell biology.

[11]  I. Mellman,et al.  Cdc42 controls secretory and endocytic transport to the basolateral plasma membrane of MDCK cells , 1999, Nature Cell Biology.

[12]  R. Scheller,et al.  The Mammalian Brain rsec6/8 Complex , 1996, Neuron.

[13]  M. Snyder,et al.  Cell polarity and morphogenesis in budding yeast. , 1998, Annual review of microbiology.

[14]  A. Bretscher,et al.  The Cooh-Terminal Domain of Myo2p, a Yeast Myosin V, Has a Direct Role in Secretory Vesicle Targeting , 1999, The Journal of cell biology.

[15]  Peter Novick,et al.  Sec3p Is a Spatial Landmark for Polarized Secretion in Budding Yeast , 1998, Cell.

[16]  D. Drubin,et al.  Origins of Cell Polarity , 1996, Cell.

[17]  M. Hall,et al.  Signaling to the actin cytoskeleton. , 1998, Annual review of cell and developmental biology.

[18]  P. Novick,et al.  Sec2p Mediates Nucleotide Exchange on Sec4p and Is Involved in Polarized Delivery of Post-Golgi Vesicles , 1997, The Journal of cell biology.

[19]  T. Pawson,et al.  A mammalian PAR-3–PAR-6 complex implicated in Cdc42/Rac1 and aPKC signalling and cell polarity , 2000, Nature Cell Biology.

[20]  M. Zerial,et al.  The diversity of Rab proteins in vesicle transport. , 1997, Current opinion in cell biology.

[21]  Zhenbiao Yang,et al.  RHO Gtpases and the Actin Cytoskeleton , 2000 .

[22]  A. Bretscher,et al.  Tropomyosin-containing actin cables direct the Myo2p-dependent polarized delivery of secretory vesicles in budding yeast. , 1998, The Journal of cell biology.

[23]  P. Novick,et al.  Protein complexes in transport vesicle targeting. , 2000, Trends in cell biology.

[24]  Samara L. Reck-Peterson,et al.  Role of actin and Myo2p in polarized secretion and growth of Saccharomyces cerevisiae. , 2000, Molecular biology of the cell.

[25]  Samara L. Reck-Peterson,et al.  The tail of a yeast class V myosin, myo2p, functions as a localization domain. , 1999, Molecular biology of the cell.

[26]  P. Novick,et al.  The role of Myo2, a yeast class V myosin, in vesicular transport , 1995, The Journal of cell biology.

[27]  R. Scheller,et al.  Sec6/8 Complex Is Recruited to Cell–Cell Contacts and Specifies Transport Vesicle Delivery to the Basal-Lateral Membrane in Epithelial Cells , 1998, Cell.

[28]  Stuart K. Kim.  Cell polarity: new PARtners for Cdc42 and Rac , 2000, Nature Cell Biology.

[29]  S. Pfeffer.  Transport-vesicle targeting: tethers before SNAREs , 1999, Nature Cell Biology.

[30]  T Watanabe,et al.  Bni1p implicated in cytoskeletal control is a putative target of Rho1p small GTP binding protein in Saccharomyces cerevisiae. , 1996, The EMBO journal.

[31]  W. Crocker,et al.  Polarity , 1910, Botanical Gazette.

[32]  P. Novick,et al.  Sec1p Binds to Snare Complexes and Concentrates at Sites of Secretion , 1999, The Journal of cell biology.

[33]  G. Joberty,et al.  The cell-polarity protein Par6 links Par3 and atypical protein kinase C to Cdc42 , 2000, Nature Cell Biology.

[34]  S. Reed,et al.  Cell cycle control of morphogenesis in budding yeast. , 1995, Current opinion in genetics & development.

[35]  R. Scheller,et al.  The sec6/8 Complex Is Located at Neurite Outgrowth and Axonal Synapse-Assembly Domains , 1999, The Journal of Neuroscience.

[36]  A. Hall,et al.  The Small GTPases Rho and Rac Are Required for the Establishment of Cadherin-dependent Cell–Cell Contacts , 1997, The Journal of cell biology.

[37]  F. Tamanoi,et al.  Rho3 of Saccharomyces cerevisiae, Which Regulates the Actin Cytoskeleton and Exocytosis, Is a GTPase Which Interacts with Myo2 and Exo70 , 1999, Molecular and Cellular Biology.

[38]  P. Novick,et al.  The exocyst is an effector for Sec4p, targeting secretory vesicles to sites of exocytosis , 1999, The EMBO journal.

[39]  P. Silver,et al.  SEC3 mutations are synthetically lethal with profilin mutations and cause defects in diploid-specific bud-site selection. , 1996, Genetics.

[40]  P. Brennwald,et al.  Sec9 is a SNAP-25-like component of a yeast SNARE complex that may be the effector of Sec4 function in exocytosis , 1994, Cell.

[41]  K. Tanaka,et al.  A downstream target of RHO1 small GTP‐binding protein is PKC1, a homolog of protein kinase C, which leads to activation of the MAP kinase cascade in Saccharomyces cerevisiae. , 1995, The EMBO journal.

[42]  E. Cabib,et al.  Role of small G proteins in yeast cell polarization and wall biosynthesis. , 1998, Annual review of biochemistry.

[43]  P. Novick,et al.  Dominant negative alleles of SEC10 reveal distinct domains involved in secretion and morphogenesis in yeast. , 1998, Molecular biology of the cell.

[44]  Y. Ohya,et al.  The Rho1 effector Pkc1, but not Bni1, mediates signalling from Tor2 to the actin cytoskeleton , 1998, Current Biology.

[45]  J. Chant,et al.  Cell polarity in yeast. , 1999, Annual review of cell and developmental biology.

[46]  J. Rothman,et al.  Mechanisms of intracellular protein transport , 1994, Nature.