A mutant plasma membrane ATPase, Pma1-10, is defective in stability at the yeast cell surface

Abstract

Pma1 is a plasma membrane H⁺-ATPase whose activity at the cell surface is essential for cell viability. In this paper we describe a temperature-sensitive pma1 allele, pma1-10 (with two point mutations in the first cytoplasmic loop of Pma1), in which the newly synthesized mutant protein fails to remain stable at the cell surface at 37°C. Instead, Pma1-10 appears to undergo internalization for vacuolar degradation in a manner dependent on End4, Vps27, Doa4, and Pep4. By contrast with wild-type Pma1, mutant Pma1-10 is hypophosphorylated and fails to associate with a Triton-insoluble fraction at 37°C, suggesting failure to enter lipid rafts. Kinetic analysis reveals that, at the permissive temperature, newly synthesized Pma1-10 acquires Triton-insolubility before becoming stabilized. We suggest that phosphorylation and lipid raft association may play important roles in maintaining protein stability at the plasma membrane.