The PXBAR-domain protein SNX9 selectively sequesters PI(3,4)P2 lipid and protects it from hydrolysis at the plasma membrane

The phosphoinositidyl lipid (PIP) family is key to spatial regulation of cellular processes, and their appearance, disappearance, and interconversion provide key signals for membrane remodeling at the plasma membrane and beyond. These signals are connected to recruitment of various peripheral membrane proteins, including sorting nexin 9 (SNX9), an important regulator of late stage endocytic and macropinocytic remodeling. Using a combination of live cell imaging, in vitro reconstitution, and atomistic molecular dynamics simulation, we probe the question of whether SNX9 is selective for PI(3,4)P2 over PI(4,5)P2, and what the physiological implications of this selectivity may be. We find that SNX9 recruitment to the plasma membrane is coincident with PI(3,4)P2 but not PI(4,5)P2 during macropinocytic ruffle maturation, and that SNX9 differentially remodels GUVs containing PI(3,4)P2 over PI(4,5)P2. Atomistic simulations further corroborate these findings and suggest that the primary mediator of this difference is a non-canonical binding interface in the fourth helix of the PX domain, which is further probed with mutational assays. We therefore hypothesize that SNX9 binding to PI(3,4)P2 over PI(4,5)P2 not only helps with modulating the spatiotemporal recruitment of SNX9 during plasma membrane remodeling processes, but also acts as a check to prevent the premature conversion of PI(3,4)P2 into PI(3)P, thus acting as a regulator of these processes.