ELIMINATION OF DEFECTIVE ALPHA-FACTOR PHEROMONE RECEPTORS

This report compares trafficking routes of a plasma membrane protein t hat was misfolded either during its synthesis or after it had reached the cell surface. A temperature-sensitive mutant form of the yeast alp ha-factor pheromone receptor (ste2-3) was found to provide a model sub strate for quality control of plasma membrane proteins. We show for th e first time that a misfolded membrane protein is recognized at the te ll surface and rapidly removed. When the ste2-3 mutant cells were cult ured continuously at 34 degrees C, the mutant receptor protein (Ste2-3 p) failed to accumulate at the plasma membrane and was degraded with a half-life of 4 min, compared with a half-life of 33 min for wild-type receptor protein (Ste2p). Degradation of both Ste2-3p and Ste2p requi red the vacuolar proteolytic activities controlled by the PEP4 gene. A t 34 degrees C, Ste2-3p comigrated with glycosylated Ste2p on sodium d odecyl sulfate-polyacrylamide gel electrophoresis, indicating that Ste 2-3p enters the secretory pathway. Degradation of Ste2-3p did not requ ire delivery to the plasma membrane as the sec1 mutation failed to blo ck rapid turnover. Truncation of the C-terminal cytoplasmic domain of the mutant receptors did not permit accumulation at the plasma membran e; thus, the endocytic signals contained ire this domain are unnecessa ry for intracellular retention. In the pep4 mutant, Ste2-3p accumulate d as series of high-molecular-weight species, suggesting a potential r ole for ubiquitin in the elimination process. When ste2-3 mutant cells were cultured continuously at 22 degrees C, Ste2-3p accumulated in th e plasma membrane. When the 22 degrees C culture was shifted to 34 deg rees C, Ste2-3p was removed from the plasma membrane and degraded by a PEP4-dependent mechanism with a 24-min half-life; the wild-type Ste2p displayed a 72-min half-life. Thus, structural defects in Ste2-3p syn thesized at 34 degrees C are recognized in transit to the plasma membr ane, leading to rapid degradation, and Ste2-3p that is preassembled at the plasma membrane is also removed and degraded following a shift to 34 degrees C.