Establishment of a robust single axis of cell polarity by coupling multiple positive feedback loops

Establishment of cell polarity—or symmetry breaking—relies on local accumulation of polarity regulators. Although simple positive feedback is sufficient to drive symmetry breaking, it is highly sensitive to stochastic fluctuations typical for living cells. Here, by integrating mathematical modelling with quantitative experimental validations, we show that in the yeast Saccharomyces cerevisiae a combination of actin- and guanine nucleotide dissociation inhibitor-dependent recycling of the central polarity regulator Cdc42 is needed to establish robust cell polarity at a single site during yeast budding. The guanine nucleotide dissociation inhibitor pathway consistently generates a single-polarization site, but requires Cdc42 to cycle rapidly between its active and inactive form, and is therefore sensitive to perturbations of the GTPase cycle. Conversely, actin-mediated recycling of Cdc42 induces robust symmetry breaking but cannot restrict polarization to a single site. Our results demonstrate how cells optimize symmetry breaking through coupling between multiple feedback loops.

[1]  Amy S. Gladfelter,et al.  Scaffold-mediated symmetry breaking by Cdc42p , 2003, Nature Cell Biology.

[2]  C. Dermardirossian,et al.  GDIs: central regulatory molecules in Rho GTPase activation. , 2005, Trends in cell biology.

[3]  C. V. Rao,et al.  Calling heads from tails: the role of mathematical modeling in understanding cell polarization. , 2009, Current opinion in cell biology.

[4]  Kay Hofmann,et al.  A positive feedback loop stabilizes the guanine‐nucleotide exchange factor Cdc24 at sites of polarization , 2002, The EMBO journal.

[5]  P. Chavrier,et al.  Cdc42 localization and cell polarity depend on membrane traffic , 2010, The Journal of cell biology.

[6]  R. Goody,et al.  RabGDI displacement by DrrA from Legionella is a consequence of its guanine nucleotide exchange activity. , 2009, Molecular cell.

[7]  Jayme M. Johnson,et al.  Symmetry breaking and the establishment of cell polarity in budding yeast. , 2011, Current opinion in genetics & development.

[8]  Herbert Waldmann,et al.  Membrane targeting mechanism of Rab GTPases elucidated by semisynthetic protein probes. , 2010, Nature chemical biology.

[9]  A. Mogilner,et al.  Cell Polarity: Quantitative Modeling as a Tool in Cell Biology , 2012, Science.

[10]  Indrani Bose,et al.  Singularity in Polarization: Rewiring Yeast Cells to Make Two Buds , 2009, Cell.

[11]  Anita T. Layton,et al.  Mechanistic mathematical model of polarity in yeast , 2012, Molecular biology of the cell.

[12]  Fabian J. Theis,et al.  MIPS: curated databases and comprehensive secondary data resources in 2010 , 2010, Nucleic Acids Res..

[13]  Jennifer M. Rust,et al.  The BioGRID Interaction Database , 2011 .

[14]  A. Wilson-Delfosse,et al.  RhoGDI-binding-defective mutant of Cdc42Hs targets to membranes and activates filopodia formation but does not cycle with the cytosol of mammalian cells. , 2001, The Biochemical journal.

[15]  J. Caviston,et al.  Singularity in budding: A role for the evolutionarily conserved small GTPase Cdc42p , 2002, Proceedings of the National Academy of Sciences of the United States of America.

[16]  T. Höfken,et al.  The Rho GDI Rdi1 regulates Rho GTPases by distinct mechanisms. , 2008, Molecular biology of the cell.

[17]  D Broek,et al.  Two types of RAS mutants that dominantly interfere with activators of RAS , 1994, Molecular and cellular biology.

[18]  J. Chant,et al.  Regulation of the polarization of T cells toward antigen-presenting cells by Ras-related GTPase CDC42. , 1995, Proceedings of the National Academy of Sciences of the United States of America.

[19]  Brian D. Slaughter,et al.  Symmetry breaking in the life cycle of the budding yeast. , 2009, Cold Spring Harbor perspectives in biology.

[20]  Anita T. Layton,et al.  Modeling Vesicle Traffic Reveals Unexpected Consequences for Cdc42p-Mediated Polarity Establishment , 2011, Current Biology.

[21]  E. Bi,et al.  Initial Polarized Bud Growth by Endocytic Recycling in the Absence of Actin Cable–dependent Vesicle Transport in Yeast , 2010, Molecular biology of the cell.

[22]  Andrew B Goryachev,et al.  Dynamics of Cdc42 network embodies a Turing‐type mechanism of yeast cell polarity , 2008, FEBS letters.

[23]  E. Bi,et al.  Central Roles of Small GTPases in the Development of Cell Polarity in Yeast and Beyond , 2007, Microbiology and Molecular Biology Reviews.

[24]  S. Emr,et al.  Vps27 recruits ESCRT machinery to endosomes during MVB sorting , 2003, The Journal of cell biology.

[25]  Timothy C. Elston,et al.  Negative Feedback Enhances Robustness in the Yeast Polarity Establishment Circuit , 2012, Cell.

[26]  S. Munro,et al.  Sorting of membrane proteins in the secretory pathway , 1993, Cell.

[27]  Jared L. Johnson,et al.  New Insights into How the Rho Guanine Nucleotide Dissociation Inhibitor Regulates the Interaction of Cdc42 with Membranes* , 2009, The Journal of Biological Chemistry.

[28]  M. Ziman,et al.  Genetic evidence for a functional interaction between Saccharomyces cerevisiae CDC24 and CDC42 , 1994, Yeast.

[29]  Douglas I. Johnson,et al.  Saccharomyces cerevisiae Cdc42p Localizes to Cellular Membranes and Clusters at Sites of Polarized Growth , 2002, Eukaryotic Cell.

[30]  Daniel J. Lew,et al.  Symmetry-Breaking Polarization Driven by a Cdc42p GEF-PAK Complex , 2008, Current Biology.

[31]  Matthias Peter,et al.  Phosphorylation of Bem2p and Bem3p may contribute to local activation of Cdc42p at bud emergence , 2007, The EMBO journal.

[32]  J. Moskow,et al.  Assembly of Scaffold-mediated Complexes Containing Cdc42p, the Exchange Factor Cdc24p, and the Effector Cla4p Required for Cell Cycle-regulated Phosphorylation of Cdc24p* , 2001, The Journal of Biological Chemistry.

[33]  Chandra L. Theesfeld,et al.  Opposing roles for actin in Cdc42p polarization. , 2005, Molecular biology of the cell.

[34]  Rong Li,et al.  Closing the loops: new insights into the role and regulation of actin during cell polarization. , 2004, Experimental cell research.

[35]  Alexander van Oudenaarden,et al.  A system of counteracting feedback loops regulates Cdc42p activity during spontaneous cell polarization. , 2005, Developmental cell.

[36]  Thomas Schmidt,et al.  Robust cell polarity is a dynamic state established by coupling transport and GTPase signaling , 2004, The Journal of cell biology.

[37]  Sigurd B. Angenent,et al.  On the spontaneous emergence of cell polarity , 2008, Nature.

[38]  S. Etienne-Manneville,et al.  Cdc42 - the centre of polarity , 2004, Journal of Cell Science.

[39]  Yoshimi Takai,et al.  Association of the Rho family small GTP-binding proteins with Rho GDP dissociation inhibitor (Rho GDI) in Saccharomyces cerevisiae , 1997, Oncogene.

[40]  Lani Wu,et al.  Spontaneous Cell Polarization Through Actomyosin-Based Delivery of the Cdc42 GTPase , 2003, Science.

[41]  Brian D. Slaughter,et al.  Non-uniform membrane diffusion enables steady-state cell polarization via vesicular trafficking , 2013, Nature Communications.

[42]  Alexandra Jilkine,et al.  A Density-Dependent Switch Drives Stochastic Clustering and Polarization of Signaling Molecules , 2011, PLoS Comput. Biol..

[43]  Brian D. Slaughter,et al.  Flippase-mediated phospholipid asymmetry promotes fast Cdc42 recycling in dynamic maintenance of cell polarity , 2012, Nature Cell Biology.

[44]  M. Snyder,et al.  Compartmentalization of the cell cortex by septins is required for maintenance of cell polarity in yeast. , 2000, Molecular cell.

[45]  Jacqueline Cherfils,et al.  RhoGDIs Revisited: Novel Roles in Rho Regulation , 2005, Traffic.

[46]  R. Cerione,et al.  RhoGDI Is Required for Cdc42-Mediated Cellular Transformation , 2003, Current Biology.

[47]  Brian D. Slaughter,et al.  Dual modes of cdc42 recycling fine-tune polarized morphogenesis. , 2009, Developmental cell.

[48]  Keith Burridge,et al.  Regulation of RhoGTPase crosstalk, degradation and activity by RhoGDI1 , 2010, Nature Cell Biology.

[49]  Yevgeny Berdichevsky,et al.  Dissociation of Rac1(GDP)·RhoGDI Complexes by the Cooperative Action of Anionic Liposomes Containing Phosphatidylinositol 3,4,5-Trisphosphate, Rac Guanine Nucleotide Exchange Factor, and GTP* , 2008, Journal of Biological Chemistry.

[50]  U. Tepass,et al.  Cdc42 and Vesicle Trafficking in Polarized Cells , 2010, Traffic.

[51]  Eugenio Marco,et al.  Endocytosis Optimizes the Dynamic Localization of Membrane Proteins that Regulate Cortical Polarity , 2007, Cell.

[52]  S. Grinstein,et al.  Phosphatidylserine is polarized and required for proper Cdc42 localization and for development of cell polarity , 2011, Nature Cell Biology.

[53]  Anthony Bretscher,et al.  Stable and dynamic axes of polarity use distinct formin isoforms in budding yeast. , 2004, Molecular biology of the cell.

[54]  Michael Knop,et al.  A versatile toolbox for PCR‐based tagging of yeast genes: new fluorescent proteins, more markers and promoter substitution cassettes , 2004, Yeast.

[55]  Aljoscha Nern,et al.  Nucleocytoplasmic Shuttling of the Cdc42p Exchange Factor Cdc24p , 2000, The Journal of cell biology.

[56]  Gary D Bader,et al.  The Genetic Landscape of a Cell , 2010, Science.

[57]  Ryohei Suzuki,et al.  Regulation of clathrin coat assembly by Eps15 homology domain–mediated interactions during endocytosis , 2012, Molecular biology of the cell.