Phase separation can regulate membrane curvature generation by bar-proteins.

Abstract

Protein phase separation enables confining protein activities in cells without requiring a membrane-enclosed compartment. Phase separation of membrane-associated proteins has been found to play central roles in cellular processes such as signaling, autophagy, and endocytosis. How phase separation regulates functionalities of membrane binding proteins is therefore a key to unravel mechanistic details of these pathways. Recently we discovered that endophilin, a member of the membrane curvature generating Bin/Amphiphysin/Rvs (BAR) protein family, undergoes liquid-liquid phase separation in association with the adapter protein lamellipodin (LPD) via multivalent interactions. Our findings suggest that endophilin-LPD phase separation plays an important role in the non-canonical endocytic pathway known as Fast Endophilin Mediated Endocytosis (FEME). We further observe that membrane curvature generation properties of endophilin are remarkably altered upon multivalent interactions with the C-terminal domain of LPD. In model membrane systems, while endophilin alone could generate tubules from negatively charged liposomes, introduction of LPD caused apparent shortening of tubule length. Investigating the membrane nanostructures with electron microscopy revealed that membrane adhesion was promoted by endo-LDP interactions on the membrane surface. Earlier, using optical microscopy, we found that LPD-mediated multivalent interactions assemble endophilin into clusters on the membrane surface. Since, the organization of endophilin on the membrane is closely related to its curvature generation functions, modulating it by a second protein could be a cellular mechanism to regulate the membrane curvature generation.