Impaired proteasome function rescues thermosensitivity of yeast cells lacking the coatomer subunit epsilon-COP

Abstract

Formation of COPI-coated transport vesicles requires a cytosolic protein complex consisting of seven subunits: alpha-, beta-, beta'-, gamma-, delta-, epsilon- and zeta-COP, collectively designated coatomer. The yeast Saccharomyces cerevisiae gene encoding the epsilon-COP subunit is known as SEC28/ANU2. anu2 null mutant cells (anu2Delta) are temperature-sensitive, and alpha-COP is rapidly degraded in these cells when they are shifted to the restrictive temperature. We isolated extragenic suppressors that rescue the temperature-sensitive growth defect of anu2Delta cells. Genetic analysis revealed that one of the suppressors is allelic to PRE8 (PRS4), which encodes a 20 S proteasome subunit. In the presence of a proteasome inhibitor, MG132, anu2Delta cells did not cease growth even at the restrictive temperature. Furthermore, MG132 inhibited the rapid decrease of alpha-COP levels in anu2Delta cells shifted to the restrictive temperature. However, secretion of certain proteins by these cells was impaired even in the presence of MG132. In conclusion, impairment of proteasome-dependent proteolysis rescued some, but not all, temperature-sensitive defects of anu2Delta cells. These results are discussed in terms of evidence that epsilon-COP plays a critical role in maintaining the structural integrity of alpha-COP.