CLATHRIN FACILITATES THE INTERNALIZATION OF 7 TRANSMEMBRANE SEGMENT RECEPTORS FOR MATING PHEROMONES IN YEAST

The role of clathrin in endocytosis of the yeast pheromone receptors w as examined using strains expressing a temperature-sensitive clathrin heavy chain. The yeast pheromone receptors belong to the family of sev en transmembrane segment, G-protein-coupled receptors. A rapid and rev ersible defect in uptake of radiolabeled alpha-factor pheromone occurr ed when the cells were transferred to the nonpermissive temperature. C onstitutive, pheromone-independent internalization of newly synthesize d a-factor pheromone receptor was also rapidly inhibited in mutant str ains at the nonpermissive temperature. In both cases residual endocyto sis, 30-50% of wild-type levels, was detected in the absence of functi onal clathrin heavy chain. Once internalized, the a-factor receptor wa s delivered to the vacuole at comparable rates in chc1-ts and wild-typ e cells at the nonpermissive temperature. Clathrin heavy chain was als o required for maximal uptake of a mutant a-factor receptor which is d ependent on pheromone for internalization. In the presence of a-factor , the internalization rate of the mutant receptor in chc1-ts cells at the nonpermissive temperature was 2.5 times slower than the rate obser ved for endocytosis of the mutant receptor in wild-type cells. These e xperiments provide in vivo evidence that clathrin plays an important r ole in the endocytosis of the seven transmembrane segment pheromone re ceptors in yeast.