ATP-dependent membrane remodeling links EHD1 functions to endocytic recycling

Raunaq Deo,Srishti Dar,Thomas J Pucadyil,Nagesh Y Kadam,Anand Srivastava,Sukrut C Kamerkar,Manish S Kushwah,Kavita Babu

doi:10.1038/s41467-018-07586-z

Abstract

Endocytic and recycling pathways generate cargo-laden transport carriers by membrane fission. Classical dynamins, which generate transport carriers during endocytosis, constrict and cause fission of membrane tubes in response to GTP hydrolysis. Relatively, less is known about the ATP-binding Eps15-homology domain-containing protein1 (EHD1), a dynamin family member that functions at the endocytic-recycling compartment. Here, we show using cross complementation assays in C. elegans that EHD1’s membrane binding and ATP hydrolysis activities are necessary for endocytic recycling. Further, we show that ATP-bound EHD1 forms membrane-active scaffolds that bulge tubular model membranes. ATP hydrolysis promotes scaffold self-assembly, causing the bulge to extend and thin down intermediate regions on the tube. On tubes below 25 nm in radius, such thinning leads to scission. Molecular dynamics simulations corroborate this scission pathway. Deletion of N-terminal residues causes defects in stable scaffolding, scission and endocytic recycling. Thus, ATP hydrolysis-dependent membrane remodeling links EHD1 functions to endocytic recycling.

Highlights

Endocytic and recycling pathways generate cargo-laden transport carriers by membrane fission
Recycling is managed by the endocytic recycling compartment (ERC) or the tubular recycling endosome (TRE), which is a dynamic organelle comprised of a network of membrane tubules and vesicles concentrated in the perinuclear region[1,2,3]
Eps15-homology domain-containing protein1 (EHD1) and 3 are localized predominantly to the ERC, EHD2 is present at the plasma membrane and EHD4 is localized to a Rab5-positive early endocytic compartment[7,8,9]

Summary

Introduction

Endocytic and recycling pathways generate cargo-laden transport carriers by membrane fission. EHD proteins contain a dynamin-like, low-affinity ATP-binding G-domain, and self-assemble into electron-dense coats on tubulated liposomes, which in turn stimulate ATP hydrolysis[8,18]. These attributes would suggest that EHDs function as classical dynamins in membrane remodeling and fission[8]. ATP hydrolysis promotes self-assembly of the scaffold, which propagates the bulge along the length of the tube causing intermediate regions to thin down and undergo fission. The N-terminal residues in EHD1 are important as their absence renders membrane bulges to catastrophically disappear in presence of ATP leading to defects in stable scaffolding, scission and endocytic recycling. We find that the closely related EHD2 shows a significantly lower ATPase activity and, concomitantly, a dramatically decreased membrane scission activity highlighting fundamental differences among EHD paralogs

Methods

Results

Conclusion