title: Membrane tethering by the atlastin GTPase depends on GTP hydrolysis but not on forming the cross-over configuration.
abstract: The membrane-anchored atlastin GTPase couples nucleotide hydrolysis to the catalysis of homotypic membrane fusion to form a branched endoplasmic reticulum network. Trans dimerization between atlastins anchored in opposing membranes, accompanied by a cross-over conformational change, is thought to draw the membranes together for fusion. Previous studies on the conformational coupling of atlastin to its GTP hydrolysis cycle have been carried out largely on atlastins lacking a membrane anchor. Consequently, whether fusion involves a discrete tethering step and, if so, the potential role of GTP hydrolysis and cross-over in tethering remain unknown. In this study, we used membrane-anchored atlastins in assays that separate tethering from fusion to dissect the requirements for each. We found that tethering depended on GTP hydrolysis, but, unlike fusion, it did not depend on cross-over. Thus GTP hydrolysis initiates stable head-domain contact in trans to tether opposing membranes, whereas cross-over formation plays a more pivotal role in powering the lipid rearrangements for fusion.
authors.affiliation: Department of Biological Sciences, Carnegie Mellon University, Pittsburgh, PA 15213.
authors.forename: Simran G
authors.initials: SG
authors.lastname: Saini
authors.affiliation: Department of Structural Biology, University of Pittsburgh, Pittsburgh, PA 15260.
authors.forename: Chuang
authors.initials: C
authors.lastname: Liu
authors.affiliation: Department of Structural Biology, University of Pittsburgh, Pittsburgh, PA 15260.
authors.forename: Peijun
authors.initials: P
authors.lastname: Zhang
authors.affiliation: Department of Biological Sciences, Carnegie Mellon University, Pittsburgh, PA 15213 thl@andrew.cmu.edu.
authors.forename: Tina H
authors.initials: TH
authors.lastname: Lee
mesh_terms: D000818:Animals; D019556:COS Cells; D002462:Cell Membrane / Q000378:metabolism* / Q000648:ultrastructure; D002522:Chlorocebus aethiops; D020285:Cryoelectron Microscopy; D029721:Drosophila Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004721:Endoplasmic Reticulum / Q000378:metabolism / Q000648:ultrastructure; D020558:GTP Phosphohydrolases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006160:Guanosine Triphosphate / Q000378:metabolism*; D006868:Hydrolysis; D008561:Membrane Fusion; D008856:Microscopy, Fluorescence; D008954:Models, Biological; D008958:Models, Molecular; D009154:Mutation; D011485:Protein Binding; D011487:Protein Conformation; D055503:Protein Multimerization; D017434:Protein Structure, Tertiary
chemical_list: D029721:Drosophila Proteins; D006160:Guanosine Triphosphate; D020558:GTP Phosphohydrolases; C543694:atl protein, Drosophila