title: Structural basis for activation, assembly and membrane binding of ESCRT-III Snf7 filaments.
abstract: The endosomal sorting complexes required for transport (ESCRTs) constitute hetero-oligomeric machines that catalyze multiple topologically similar membrane-remodeling processes. Although ESCRT-III subunits polymerize into spirals, how individual ESCRT-III subunits are activated and assembled together into a membrane-deforming filament remains unknown. Here, we determine X-ray crystal structures of the most abundant ESCRT-III subunit Snf7 in its active conformation. Using pulsed dipolar electron spin resonance spectroscopy (PDS), we show that Snf7 activation requires a prominent conformational rearrangement to expose protein-membrane and protein-protein interfaces. This promotes the assembly of Snf7 arrays with ~30 Å periodicity into a membrane-sculpting filament. Using a combination of biochemical and genetic approaches, both in vitro and in vivo, we demonstrate that mutations on these protein interfaces halt Snf7 assembly and block ESCRT function. The architecture of the activated and membrane-bound Snf7 polymer provides crucial insights into the spatially unique ESCRT-III-mediated membrane remodeling.
authors.affiliation: Weill Institute of Cell and Molecular Biology, Cornell University, Ithaca, United States.|Department of Molecular Biology and Genetics, Cornell University, Ithaca, United States.
authors.forename: Shaogeng
authors.initials: S
authors.lastname: Tang
authors.affiliation: Weill Institute of Cell and Molecular Biology, Cornell University, Ithaca, United States.|Department of Molecular Biology and Genetics, Cornell University, Ithaca, United States.
authors.forename: W Mike
authors.initials: WM
authors.lastname: Henne
authors.affiliation: National Biomedical Center for Advanced Electron Spin Resonance Technology, Cornell University, Ithaca, United States.|Department of Chemistry and Chemical Biology, Cornell University, Ithaca, United States.
authors.forename: Peter P
authors.initials: PP
authors.lastname: Borbat
authors.affiliation: Weill Institute of Cell and Molecular Biology, Cornell University, Ithaca, United States.|Department of Molecular Biology and Genetics, Cornell University, Ithaca, United States.
authors.forename: Nicholas J
authors.initials: NJ
authors.lastname: Buchkovich
authors.affiliation: National Biomedical Center for Advanced Electron Spin Resonance Technology, Cornell University, Ithaca, United States.|Department of Chemistry and Chemical Biology, Cornell University, Ithaca, United States.
authors.forename: Jack H
authors.initials: JH
authors.lastname: Freed
authors.affiliation: Weill Institute of Cell and Molecular Biology, Cornell University, Ithaca, United States.|Department of Molecular Biology and Genetics, Cornell University, Ithaca, United States.
authors.forename: Yuxin
authors.initials: Y
authors.lastname: Mao
authors.affiliation: Weill Institute of Cell and Molecular Biology, Cornell University, Ithaca, United States.|Department of Molecular Biology and Genetics, Cornell University, Ithaca, United States.
authors.forename: J Christopher
authors.initials: JC
authors.lastname: Fromme
authors.affiliation: Weill Institute of Cell and Molecular Biology, Cornell University, Ithaca, United States.|Department of Molecular Biology and Genetics, Cornell University, Ithaca, United States.
authors.forename: Scott D
authors.initials: SD
authors.lastname: Emr
mesh_terms: D018360:Crystallography, X-Ray; D056827:Endosomal Sorting Complexes Required for Transport / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D011992:Endosomes / Q000378:metabolism*; D008566:Membranes / Q000378:metabolism*; D008958:Models, Molecular; D050505:Mutant Proteins / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D011487:Protein Conformation; D055503:Protein Multimerization; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D013057:Spectrum Analysis
chemical_list: D056827:Endosomal Sorting Complexes Required for Transport; D050505:Mutant Proteins; C083890:SNF7 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins
keywords: ESCRT-III; PDS spectroscopy; S. cerevisiae; Snf7; X-ray crystal structure; biophysics; cell biology; protofilament; structural biology; structural rearrangement