Characterization of the Wsc1 Protein, a Putative Receptor in the Stress Response of Saccharomyces cerevisiae

Corresponding author: Roymarie Ballester, Department of Molecular, Cellular and Developmental Biology, University of California, Santa Barbara, CA 93106., balleste{at}lifesci.ucsb.edu (E-mail)

Communicating editor: F. WINSTON

Wsc1p, Wsc2p, Wsc3p, and Wsc4p, members of a novel protein family in the yeast Saccharomyces cerevisiae, are putative sensors or receptors in the stress response. Genetic characterization suggests that the WSC family are upstream regulators of the stress-activated PKC1-MAP kinase cascade and are required for the heat shock response and for maintenance of cell wall integrity. The Wsc proteins share sequence characteristics: at their N terminus they have a cysteine motif and a serine/threonine-rich domain predicted to be extracellular, a hydrophobic domain suggested to be transmembranous, and a variable, highly charged C terminus that may be involved in intracellular signaling. Although a role for the WSC genes in maintenance of cell wall integrity has been firmly established, little is known about the properties of the proteins. As reported here, to study its properties in vivo, we epitope tagged the Wsc1 protein. Wsc1p was found to fractionate with the membrane pellet after high-speed centrifugation. Extraction experiments show that Wsc1p is an integral membrane protein present in two forms: one solubilized by detergent, the other Triton X-100 insoluble. Our results also show that Wsc1p is glycosylated and phosphorylated. To characterize the contribution of different domains to the function of Wsc1p, we generated various deletion constructs. Analysis of the properties and function of the mutant proteins shows that the predicted extracellular serine/threonine-rich domain is required for Wsc1p functionality, as well as its glycosylation. A mutant Wsc1 protein lacking the putative transmembrane domain is not functional and partitions to the soluble fraction. Overexpression of full-length Wsc1p inhibits cell growth, with the N terminus alone being sufficient for this inhibition. This suggests that Wsc1p may function in a complex with at least one other protein important for normal cell growth.

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