SGIP1α induces membrane tubulation and forms homodimer via its unique additional hydrophobic region in MP domain

Abstract

The early steps of clathrin-mediated endocytosis (CME) require membrane deformation and invaginations, and BAR domains or FCH domain proteins such as endophilin or PACSIN have been implicated in these processes. SGIP1 (SH3-domain GRB2-like endophilin interacting protein 1) is an endophilin interacting protein and is composed of N-terminal membrane phospholipid binding (MP) domain, proline rich domain (PRD) in the middle, and C-terminal µ-homology domain (µHD). It is known to be specifically expressed in the brain and to function in the regulation of energy homeostasis although its role in the nervous system is not fully understood. Recent study showed that its the longest splicing variant, SGIP1α, plays a role in CME by interacting with phospholipids and Eps15 and that MP domain of SGIP1α binds to phospholipid and deforms the plasma membrane. The MP domain, however, doesnt shows any sequence homology to well-known tubule-forming BAR or EFC domain, therefore how MP domain of SGIP1α induces membrane tubulation has been in question. Here, I found that unlike SGIP1, SGIP1α has additional 28 amino acids in MP domain. I also found that the additional 28 amino acids in MP domain are essential for SGIP1α to induce membrane tubule while SGIP1 which lacks this additional amino acids failed to induce membrane tubule. The sequence analysis of additional 28 amino acids in MP domain identified a highly hydrophobic region, flanked by two positively charged regions. Using various point mutants of MP domain, I found that the upstream of hydrophobic region of SGIP1α is crucial for tubulation. I also found that SGIP1α forms homodimer via MP domain and the region between the hydrophobic region and the subsequent positively charged region is responsible for homodimerization. Evidently, SGIP1α mutants lacking hydrophobic region forms neither membrane tubule nor homodimer. Taken together, my results suggest that the hydrophobic region in additional 28 amino acids of SGIP1α MP domain plays an essential role in membrane tubulation and homodimerization. Considering that SGIP1 is a brain-specific protein, the unique feature in SGIP1α MP domain may confer its special role in nervous system although it requires further study.