Adaptor complex-independent clathrin function in yeast

Clathrin-associated adaptor protein (AP) complexes are major structural com ponents of clathrin-coated vesicles, functioning in clathrin coat assembly and cargo selection. We have carried out a systematic biochemical and genet ic characterization of AP complexes in Saccharomyces cerevisiae. Using coim munoprecipitation, the subunit composition of two complexes, AP-1 and AP-2R , has been defined. These results allow assignment of the 13 potential AP s ubunits encoded in the yeast genome to three AP complexes. As assessed by i n vitro binding assays and coimmunoprecipitation, only AP-1 interacts with clathrin. Individual or combined disruption of AP-1 subunit genes in cells expressing a temperature-sensitive clathrin heavy chain results in accentua ted growth and cy-factor pheromone maturation defects, providing further ev idence that AP-1 is a clathrin adaptor complex. However, in cells expressin g wild-type clathrin, the same AP subunit deletions have no effect on growt h or ct-factor maturation. Furthermore, gel filtration chromatography revea led normal elution patterns of clathrin-coated vesicles in cells lacking AP -1. Similarly, combined deletion of genes encoding the beta subunits of the three AP complexes did not produce defects in clathrin-dependent sorting i n the endocytic and vacuolar pathways or alterations in gel filtration prof iles of clathrin-coated vesicles. We conclude that AP complexes are dispens able for clathrin function in S, cerevisiae under normal conditions. Our re sults suggest that alternative factors assume key roles in stimulating clat hrin coat assembly and cargo selection during clathrin-mediated vesicle for mation in yeast.