DNM1, A DYNAMIN-RELATED GENE, PARTICIPATES IN ENDOSOMAL TRAFFICKING IN YEAST

We identified DNM1, a novel dynamin-related gene in Saccharomyces cere visiae. Molecular and genetic mapping showed that DNM1 is the most pro ximal gene to the right of centromere 12, and is predicted to encode a protein of 85 kD, designated Dnm1p. The protein exhibits 41% overall identity with full-length dynamin I and 55% identity with the most hig hly conserved 400-amino acid GTPase region. Our findings show that lik e mammalian dynamin, Dnm1p participates in endocytosis; however, it is unlikely to be a cognate homologue. Cells with a disruption in the DN M1 gene showed mating response defects consistent with a delay in rece ptor-mediated endocytosis. The half-life of the Ste3p pheromone recept or was increased two- to threefold in the dnm1 mutant, demonstrating t hat Dnm1p participates in the constitutive turnover of the receptor. T o define the step in the endocytic pathway at which Dnm1p acts, we ana lyzed mutant strains at both early and late steps of the process. Init ial internalization of epitope-tagged pheromone receptor or of labeled pheromone proceeded with wild-type kinetics. However, delivery of the internalized receptor to the vacuole was greatly impeded during ligan d-induced endocytosis. These data suggest that during receptor-mediate d endocytosis, Dnm1p acts after internalization, but before fusion wit h the vacuole. The dnm1 mutant was not defective for sorting of vacuol ar proteins, indicating that Dnm1p is not required for transport from the late endosome to the vacuole. Therefore, we suggest that Dnm1p par ticipates at a novel step before fusion with the late endosome.