The auxilin-like phosphoprotein Swa2p is required for clathrin function inyeast

Background: In eukaryotic cells, clathrin-coated Vesicles transport specifi c cargo from the plasma membrane and trans-Golgi network to the endosomal s ystem. Removal of the clathrin coat in vitro requires the uncoating ATPase Hsc70 and its DnaJ cofactor auxilin. To date, a requirement for auxilin and Hsc70 in clathrin function in vivo has not been demonstrated. Results: The Saccharomyces cerevisiae SWAP gene, previously identified in a synthetic lethal screen with arf1, was cloned and found to encode a protei n with a carboxy-terminal DnaJ domain which is homologous to that of auxili n. Like auxilin, Swa2p has a clathrin-binding domain and is able to stimula te the ATPase activity of Hsc70. The swa2-1 allele recovered from the origi nal screen carries a point mutation in its tetratricopeptide repeat (TPR) d omain, a motif not found in auxilin but known in other proteins to mediate interaction with heat-shock proteins. Swa2p fractionates in the cytosol and appears to be heavily phosphorylated. Disruption of SWAP causes slow growt h and several phenotypes that are very similar to those exhibited by clathr in mutants. Furthermore, the swa2 Delta mutant exhibits a significant incre ase in membrane-associated or -assembled clathrin relative to a wild-type s train. Conclusions: These results indicate that Swa2p is a clathrin-binding protei n required for normal clathrin function in vivo. They suggest that Swa2p is the yeast ortholog of auxilin and has a role in disassembling clathrin, no t only in uncoating clathrin-coated vesicles but perhaps in preventing unpr oductive clathrin assembly in vivo.