Mutational analysis of the cytoplasmic domain of the Wsc1 cell wall stress sensor.

Wsc1 is a member of a family of highly O-glycosylated cell surface proteins that reside in the plasma membrane of Saccharomyces cerevisiae and function as sensors of cell wall stress. These proteins activate the cell wall integrity signalling pathway by stimulating the small G-protein Rho1, protein kinase C (Pkc1) and a MAP kinase cascade. The cytoplasmic domains of Wsc1 family members interact with the Rom2 guanine nucleotide exchange factor to stimulate GTP-binding of Rho1. Here, a mutational analysis of the cytoplasmic domain of Wsc1 is presented. The data identify two regions of the Wsc1 cytoplasmic tail that are conserved with other family members as important for Rom2 interaction. These regions are separated by an inhibitory region, which includes a cluster of seryl residues that appear to be phosphorylated. Mutational analysis of these residues supports a model in which Wsc1 interaction with Rom2 is negatively regulated by phosphorylation.

[1]  U. Jung,et al.  Regulation of the yeast Rlm1 transcription factor by the Mpk1 cell wall integrity MAP kinase , 2002, Molecular microbiology.

[2]  D. E. Levin,et al.  Wsc1 and Mid2 Are Cell Surface Sensors for Cell Wall Integrity Signaling That Act through Rom2, a Guanine Nucleotide Exchange Factor for Rho1 , 2001, Molecular and Cellular Biology.

[3]  M. Molina,et al.  Regulatory Mechanisms for Modulation of Signaling through the Cell Integrity Slt2-mediated Pathway in Saccharomyces cerevisiae * , 2000, The Journal of Biological Chemistry.

[4]  R. Ballester,et al.  Characterization of the Wsc1 protein, a putative receptor in the stress response of Saccharomyces cerevisiae. , 1999, Genetics.

[5]  B. Errede,et al.  Mid2 Is a Putative Sensor for Cell Integrity Signaling in Saccharomyces cerevisiae , 1999, Molecular and Cellular Biology.

[6]  Troy Ketela,et al.  Saccharomyces cerevisiae Mid2p Is a Potential Cell Wall Stress Sensor and Upstream Activator of thePKC1-MPK1 Cell Integrity Pathway , 1999, Journal of bacteriology.

[7]  J. Heinisch,et al.  A screen for upstream components of the yeast protein kinase C signal transduction pathway identifies the product of the SLG1 gene , 1998, Molecular and General Genetics MGG.

[8]  R. Ballester,et al.  A family of genes required for maintenance of cell wall integrity and for the stress response in Saccharomyces cerevisiae. , 1997, Proceedings of the National Academy of Sciences of the United States of America.

[9]  B. Errede,et al.  Coordination of the mating and cell integrity mitogen-activated protein kinase pathways in Saccharomyces cerevisiae , 1997, Molecular and cellular biology.

[10]  I. Herskowitz,et al.  A role for the Pkc1 MAP kinase pathway of Saccharomyces cerevisiae in bud emergence and identification of a putative upstream regulator , 1997, The EMBO journal.

[11]  Y. Watanabe,et al.  Characterization of a serum response factor-like protein in Saccharomyces cerevisiae, Rlm1, which has transcriptional activity regulated by the Mpk1 (Slt2) mitogen-activated protein kinase pathway , 1997, Molecular and cellular biology.

[12]  Marc Bickle,et al.  The Yeast Phosphatidylinositol Kinase Homolog TOR2 Activates RHO1 and RHO2 via the Exchange Factor ROM2 , 1997, Cell.

[13]  K Tanaka,et al.  Rom1p and Rom2p are GDP/GTP exchange proteins (GEPs) for the Rho1p small GTP binding protein in Saccharomyces cerevisiae. , 1996, The EMBO journal.

[14]  Y. Anraku,et al.  Activation of Yeast Protein Kinase C by Rho1 GTPase (*) , 1996, The Journal of Biological Chemistry.

[15]  T Watanabe,et al.  Identification of Yeast Rho1p GTPase as a Regulatory Subunit of 1,3-β-Glucan Synthase , 1996, Science.

[16]  Guang-Chao Chen,et al.  Rho1p, a Yeast Protein at the Interface Between Cell Polarization and Morphogenesis , 1996, Science.

[17]  D. E. Levin,et al.  A Second Osmosensing Signal Transduction Pathway in Yeast , 1995, The Journal of Biological Chemistry.

[18]  K. Irie,et al.  Dynamics and organization of MAP kinase signal pathways , 1995, Molecular reproduction and development.

[19]  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.

[20]  N. Morin,et al.  Differential expression and function of two homologous subunits of yeast 1,3-beta-D-glucan synthase , 1995, Molecular and cellular biology.

[21]  M. Snyder,et al.  Molecular basis of cell integrity and morphogenesis in Saccharomyces cerevisiae. , 1995, Microbiological reviews.

[22]  T. Watanabe,et al.  Characterization and gene cloning of 1,3-beta-D-glucan synthase from Saccharomyces cerevisiae. , 1995, European journal of biochemistry.

[23]  U. Jung,et al.  The protein kinase C-activated MAP kinase pathway of Saccharomyces cerevisiae mediates a novel aspect of the heat shock response. , 1995, Genes & development.

[24]  F. Klis,et al.  Identification of two cell cycle regulated genes affecting the β1,3‐glucan content of cell walls in Saccharomyces cerevisiae , 1995, FEBS letters.

[25]  N. Morin,et al.  The Saccharomyces cerevisiae FKS1 (ETG1) gene encodes an integral membrane protein which is a subunit of 1,3-beta-D-glucan synthase. , 1994, Proceedings of the National Academy of Sciences of the United States of America.

[26]  Clarence S. M. Chan,et al.  Control of cellular morphogenesis by the Ip12/Bem2 GTPase-activating protein: possible role of protein phosphorylation , 1994, The Journal of cell biology.

[27]  F. Klis Review: Cell wall assembly in yeast , 1994, Yeast.

[28]  K. Irie,et al.  MKK1 and MKK2, which encode Saccharomyces cerevisiae mitogen-activated protein kinase-kinase homologs, function in the pathway mediated by protein kinase C , 1993, Molecular and cellular biology.

[29]  Y. Watanabe,et al.  A yeast mitogen-activated protein kinase homolog (Mpk1p) mediates signalling by protein kinase C , 1993, Molecular and cellular biology.

[30]  J. Carpentier,et al.  The osmotic integrity of the yeast cell requires a functional PKC1 gene product , 1992, Molecular and cellular biology.

[31]  D. E. Levin,et al.  Mutants in the S. cerevisiae PKC1 gene display a cell cycle-specific osmotic stability defect , 1992, The Journal of cell biology.

[32]  M. Snyder,et al.  A synthetic lethal screen identifies SLK1, a novel protein kinase homolog implicated in yeast cell morphogenesis and cell growth , 1992, Molecular and cellular biology.

[33]  D. E. Levin,et al.  Dominant mutations in a gene encoding a putative protein kinase (BCK1) bypass the requirement for a Saccharomyces cerevisiae protein kinase C homolog , 1992, Molecular and cellular biology.

[34]  M. Molina,et al.  A protein kinase gene complements the lytic phenotype of Saccharomyces cerevisiae lyt2 mutants , 1991, Molecular microbiology.

[35]  D. E. Levin,et al.  A candidate protein kinase C gene, PKC1, is required for the S. cerevisiae cell cycle , 1990, Cell.

[36]  R. Sikorski,et al.  A system of shuttle vectors and yeast host strains designed for efficient manipulation of DNA in Saccharomyces cerevisiae. , 1989, Genetics.

[37]  S. Ho,et al.  Site-directed mutagenesis by overlap extension using the polymerase chain reaction. , 1989, Gene.

[38]  K. Murata,et al.  Transformation of intact yeast cells treated with alkali cations , 1983 .

[39]  G. Fink,et al.  Methods in yeast genetics , 1979 .