Sequences Flanking The Transmembrane Segments Facilitate Mitochondrial Localization And Membrane Fusion By Mitofusin

Mitochondria are double membrane-bound organelles involved in many critical processes,including ATP synthesis and cell apoptotic.Mitochondria are highly dynaminc with constantly divide and fuse to maintain its proper morphology and function.The mitochondrial fusion needs outer membrane fusion and inner membrane fusion seperately.Homotypic fusion of the outer mitochondrial membranes requires the mitofusin(MFN)proteins,a family of dynamin-like GTPases.MFNs are anchored in the membrane by transmembrane(TM)segments,exposing both the N-terminal GTPase domain,Helix Bundle(HB)and the C-terminal tail(CT)to the cytosol.This arrangement is very similar to that of the atlastin(ATL)GTPases,which mediate fusion of endoplasmic reticulum(ER)membranes.ATL is also a membrane protein belonging to the dynamin family.Based on several different crystal structures of the cytosolic domain of ATL and biochemical assayes,fusion model of ATL has been prososed.Given that MFN is difficulit to purify,its fusion mechanism is poorly understood.Even whether MFN alone is sufficient to mediate membrane fusion is still unclear.In order to gain insights into the working mechanism and domains' function of MFN,we engineered various MFN-ATL chimeras to investigate MFN-mediated fusion.When MFN1 is localized to the ER by TM swapping with ATL1,it functions in the maintenance of ER morphology and fusion.In addition,an amphipathic helix a10 in the CT of MFN1 is exchangeable with that of ATL1 and critical for mitochondrial localization of MFN1.Furthermore,hydrophobic residues in a7a8 loop also play a role in membrane targeting but not fusion.Our findings provide important insight into MFN-mediated membrane fusion and mitochondrial localization.