A novel EH domain protein of Saccharomyces cerevisiae, Ede1p, involved in endocytosis

Sequencing of the entire genome of S, cerevisiae has revealed the existence of five proteins containing EH domains, These are protein-protein interact ion modules first described in mammalian Eps15, a protein that is involved in clathrin-dependent endocytosis. Two of the yeast proteins, End3p and Pan 1p, are required for the internalization step of endocytosis, me report cha racterization of the nonessential ORF YBL047c which, like Eps15, encodes a protein with three N-terminal EH domains. Deletion of YBL047c leads to a de fective fluid-phase endocytosis and to defective internalization of the phe romone a-factor and uracil permease, We therefore named YBL047c EDE1, for E H Domains and Endocytosis. Ede1p expressed as a chromosomally encoded fusio n to the green fluorescent protein is localized in punctate cortical spots that only partially colocalize with actin patches. This localization is mai ntained when actin is depolymerized, Deletion of EDE1 impairs the diploid b udding pattern, but has only a small impact on actin cytoskeleton organizat ion, in contrast to the effects observed in pan1 cells and many end mutants impaired in proteins colocalizing with cortical actin patches. Genetic int eraction was observed between EDE1 and RSP5, which encodes the ubiquitin li gase Rsp5p essential for ubiquitin-dependent endocytosis of many plasma mem brane proteins, thus further emphasizing the functional link between Rsp5p and the EH domain proteins. We also observed genetic interaction between ED E1, and END3 or PAN1, suggesting that Ede1p might be part of a yeast EH net work implicated in endocytosis.