Identifying New Endocytosis Components

One highly successful focus has been our group’s comprehensive identification of novel proteins that control endocytic transport, and our subsequent dissection of their mechanisms of action. Using the tagged ligand assay system that we invented following yolk protein YP170, our laboratory systematically tested nearly every gene in the worm genome by RNAi for effects on the basolateral secretion of yolk proteins by the intestinal epithelium and its subsequent endocytosis by the oocytes (Balklava et al., 2007). After secondary assays our studies identified hundreds of endocytosis and secretion regulators, many of which are highly conserved among metazoans (including humans) but are not found in simpler systems like yeast (Sato et al., 2005; Sato et al., 2008). One broad conclusion we drew from these screens was the critical importance of endocytic recycling, which must occur to support endocytic uptake. Deep analysis of our mutant collection showed that many of the novel trafficking regulators we discovered controlled receptor recycling steps rather than directly affecting uptake. Our data were pivotal in defining endocytic recycling in general as an active process, rather than a passive default pathway, and also served as motivation for our group to move on to deciphering recycling mechanisms, which were extremely poorly defined across all systems.