Poster Presentation 2014 International Biophysics Congress

Molecular mechanism for atlastin-mediated homotypic fusion of endoplasmic reticulum membranes (#305)

Xin Bian 1 , Tina Liu 2 3 , Sha Sun 1 , Tom Rapoport 2 3 , Junjie Hu 1
  1. Nankai University, Tianjin, China
  2. Department of Cell Biology, Harvard Medical School, Boston, MA, USA
  3. Howard Hughes Medical Institute, Boston, MA, USA

The homotypic fusion of endoplasmic reticulum (ER), which is required for ER network formation, is catalyzed by the dynamin-like, membrane-bound GTPase atlastin (ATL). ATL consists of an N-terminal cytosolic domain, containing a GTPase module and a three-helix bundle (3HB), followed by two transmembrane segments (TMs) and a C-terminal tail (CT). Here, we have addressed the mechanism of ATL-mediated membrane fusion. The crystal structure of the cytosolic domain of human ATL1 was determined in the presence of GDP. Interestingly, we obtained two different structures. This conformational change and biochemical analyses demonstrate that, during fusion, the initial contact between ATL molecules in opposing membranes is mediated by the GTPase domains, which form a dimer with the nucleotides bound at the interface. Upon GTP hydrolysis and phosphate release, the 3HBs of the two ATL molecules undergo a major conformational change relative to the GTPase domains, which pulls the membranes together. The CT and TMs are also required for fusion. The CT can form an amphipathic helix and facilitates vesicle fusion by directly interacting with and perturbing the lipid bilayer. A synthetic peptide corresponding to the helix can act in trans to restore the fusion activity of tailless ATL. The TMs are more than just membrane anchors, because they cannot be deleted, mutated or replaced with unrelated TMs. Coimmunoprecipitation experimants prove that TMs can mediate the nucleotide-independent oligomerization of ATL to promote membrane fusion. Finally, we also found at early stage of fusion reaction, continuous GTP hydrolysis is required for ATL to tether vesicles. The linker region between the 3HB and the TMs is important for the transition from tethered state to fusion. Taken together, we propose a model in which different domains of ATL cooperate to mediate membrane fusion. Our results also provide some explanations regarding how ATL1 mutations cause hereditary spastic paraplegia.