Endocytic Sorting and Recycling Require Membrane Phosphatidylserine Asymmetry Maintained by TAT-1/CHAT-1

Abstract

Endocytic sorting is achieved through the formation of morphologically and functionally distinct sub-domains within early endosomes. Cargoes destined for recycling are sorted to and transported through newly-formed tubular membranes, but the processes that regulate membrane tubulation are poorly understood. Here, we identified a novel Caenorhabditis elegans Cdc50 family protein, CHAT-1, which acts as the chaperone of the TAT-1 P4-ATPase to regulate membrane phosphatidylserine (PS) asymmetry and endocytic transport. In chat-1 and tat-1 mutants, the endocytic sorting process is disrupted, leading to defects in both cargo recycling and degradation. TAT-1 and CHAT-1 colocalize to the tubular domain of the early endosome, the tubular endocytic recycling compartment (ERC), and the recycling endosome where PS is enriched on the cytosolic surface. Loss of tat-1 and chat-1 function disrupts membrane PS asymmetry and abrogates the tubular membrane structure. Our data suggest that CHAT-1 and TAT-1 maintain membrane phosphatidylserine asymmetry, thus promoting membrane tubulation and regulating endocytic sorting and recycling.

Author Summary

The process by which cells take up nutrients and other large molecules from the extracellular environment is known as endocytosis. At the cell surface, external molecules become enclosed in membrane spheres called endosomes. Early endosomes serve as a sorting station, directing the contents (cargo molecules) to the correct compartment within the cell. This is thought to be achieved by the formation of membrane structures with distinct shape and function. For example, cargoes destined for recycling and degradation are processed through tubular membrane structures and big vesicular compartments, respectively. However, it is poorly understood how early endosome membranes are shaped into different structures. Here we show that two proteins, CHAT-1 and TAT-1, regulate membrane structure and are important for normal endocytic transport in the nematode worm C. elegans. TAT-1 and CHAT-1 are found in tubular membrane structures along the sorting and recycling pathway, where they enrich the outer membrane layer with a lipid called phosphatidylserine (PS) and probably change the membrane curvature. Loss of tat-1 and chat-1 function disrupts the asymmetric distribution of PS, abolishes tubular membrane structures, and abrogates endocytic sorting/recycling. Our data support a role of TAT-1/CHAT-1–regulated membrane PS asymmetry in promoting membrane tubulation for endocytic cargo sorting and recycling.