AMPH-1/Amphiphysin/Bin1 functions with RME-1/Ehd1 in endocytic recycling

Abstract

The RME1 ATPases are implicated in endocytic recycling. C. elegans RME1 interacts with Amphiphysin to regulate endocytic recycling in vivo and the two proteins cooperate in the generation of cargo carriers in vitro. The interaction is conserved in other eukaryotes. RME-1/EHD1 (receptor mediated endocytosis/Eps15 homology-domain containing 1) family proteins are key residents of the recycling endosome, which are required for endosome-to-plasma membrane transport in Caenorhabditis elegans and mammals. Recent studies suggest similarities between the RME-1/EHD proteins and the Dynamin GTPase superfamily of mechanochemical pinchases, which promote membrane fission. Here we show that endogenous C. elegans AMPH-1, the only C. elegans member of the Amphiphysin/BIN1 family of BAR (Bin1-Amphiphysin-Rvs161p/167p)-domain-containing proteins, colocalizes with RME-1 on recycling endosomes in vivo, that amph-1-deletion mutants are defective in recycling endosome morphology and function, and that binding of AMPH-1 Asn-Pro-Phe(Asp/Glu) sequences to the RME-1 EH-domain promotes the recycling of transmembrane cargo. We also show a requirement for human BIN1 (also known as Amphiphysin 2) in EHD1-regulated endocytic recycling. In vitro, we find that purified recombinant AMPH-1–RME-1 complexes produce short, coated membrane tubules that are qualitatively distinct from those produced by either protein alone. Our results indicate that AMPH-1 and RME-1 cooperatively regulate endocytic recycling, probably through functions required for the production of cargo carriers that exit the recycling endosome for the cell surface.