A synthetic analysis of the Saccharomyces cerevisiae stress sensor Mid2p, and identification of a Mid2p-interacting protein, Zeo1p, that modulates the PKC1-MPK1 cell integrity pathway.
Mid2p is a plasma membrane protein that functions in Saccharomyces cerevisiae as a sensor of cell wall stress, activating the PKC1-MPK1 cell integrity pathway via the small GTPase Rho1p during exposure to mating pheromone, calcofluor white, and heat. To examine Mid2p signalling, a global synthetic interaction analysis of a mid2 mutant was performed; this identified 11 interacting genes. These include WSC1 and ROM2, upstream elements in cell integrity pathway signalling, and FKS1 and SMI1, required for 1,3-beta-glucan synthesis. These synthetic interactions indicate that the Wsc1p sensor acts through Rom2p to activate the Fks1p glucan synthase in a Mid2p-independent way. To further explore Mid2p signalling a two-hybrid screen was done using the cytoplasmic tail of Mid2p; this identified ZEO1 (YOL109w), encoding a 12 kDa peripheral membrane protein that localizes to the plasma membrane. Disruption of ZEO1 leads to resistance to calcofluor white and to a Mid2p-dependent constitutive phosphorylation of Mpk1p, supporting a role for Zeo1p in the cell integrity pathway. Consistent with this, zeo1-deficient cells suppress the growth defect of mutants in the Rho1p GDP-GTP exchange factor Rom2p, while exacerbating the growth defect of sac7delta mutants at 37 degrees C. In contrast, mid2delta mutants have opposing effects to zeo1delta mutants, being synthetically lethal with rom2delta, and suppressing an 18 degrees C growth defect of sac7delta, while overexpression of MID2 rescues a rom2delta 37 degrees C growth defect. Thus, MID2 and ZEO1 appear to play reciprocal roles in the modulation of the yeast PKC1-MPK1 cell integrity pathway.
Green
Robin
R
Department of Biology, McGill University, Montreal, Quebec, Canada H3A 1B1.
Lesage
Guillaume
G
Department of Biology, McGill University, Montreal, Quebec, Canada H3A 1B1.
Sdicu
Anne-Marie
AM
Department of Biology, McGill University, Montreal, Quebec, Canada H3A 1B1.
Ménard
Patrice
P
Department of Biology, McGill University, Montreal, Quebec, Canada H3A 1B1.
Bussey
Howard
H
Department of Biology, McGill University, Montreal, Quebec, Canada H3A 1B1.
D048868:Adaptor Proteins, Signal Transducing; D002135:Calcium-Binding Proteins / Q000378:metabolism*; D047908:Intracellular Signaling Peptides and Proteins; D008562:Membrane Glycoproteins; D008565:Membrane Proteins / Q000378:metabolism*; D020928:Mitogen-Activated Protein Kinases; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism*; D015398:Signal Transduction
D048868:Adaptor Proteins, Signal Transducing; D002135:Calcium-Binding Proteins; D047908:Intracellular Signaling Peptides and Proteins; C091625:MID2 protein, S cerevisiae; D008562:Membrane Glycoproteins; D008565:Membrane Proteins; D029701:Saccharomyces cerevisiae Proteins; C482121:Zeo1 protein, S cerevisiae; D020928:Mitogen-Activated Protein Kinases; C072724:SLT2 protein, S cerevisiae