PT  - JOURNAL ARTICLE
AU  - Mateusz Kozak
AU  - Marko Kaksonen
TI  - Phase separation of Ede1 promotes the initiation of endocytic events
AID  - 10.1101/861203
DP  - 2019 Jan 01
TA  - bioRxiv
PG  - 861203
4099  - http://biorxiv.org/content/early/2019/12/02/861203.short
4100  - http://biorxiv.org/content/early/2019/12/02/861203.full
AB  - Clathrin-mediated endocytosis is a major pathway that eukaryotic cells use to produce transport vesicles from the plasma membrane. The assembly of the endocytic coat is initiated by a dynamic network of weakly interacting proteins, but the exact mechanism of initiation is unknown. Ede1, the yeast homologue of mammalian Eps15, is one of the early-arriving endocytic proteins and a key initiation factor. In the absence of Ede1, most other early endocytic proteins lose their punctate localization and the frequency of endocytic initiation is decreased. We show here that in mutants with increased amounts of cytoplasmic Ede1, the excess protein forms large condensates which exhibit properties of phase separated liquid protein droplets. These Ede1 condensates recruit many other early-arriving endocytic proteins. Their formation depends on the core region of Ede1 that contains a coiled coil and a low-complexity domain. We demonstrate that Ede1 core region is essential for the endocytic function of Ede1. The core region can also promote clustering of a heterologous lipid-binding domain into discrete sites on the plasma membrane that initiate endocytic events. We propose that the clustering of the early endocytic proteins and cargo depend on phase separation mediated by Ede1.