The endosomal sorting complex ESCRT-II mediates the assembly and architecture of ESCRT-III helices.
The endosomal sorting complexes required for transport (ESCRTs) constitute hetero-oligomeric machines that mediate topologically similar membrane-sculpting processes, including cytokinesis, retroviral egress, and multivesicular body (MVB) biogenesis. Although ESCRT-III drives membrane remodeling that creates MVBs, its structure and the mechanism of vesicle formation are unclear. Using electron microscopy, we visualize an ESCRT-II:ESCRT-III supercomplex and propose how it mediates vesicle formation. We define conformational changes that activate ESCRT-III subunit Snf7 and show that it assembles into spiraling ~9 nm protofilaments on lipid monolayers. A high-content flow cytometry assay further demonstrates that mutations halting ESCRT-III assembly block ESCRT function. Strikingly, the addition of Vps24 and Vps2 transforms flat Snf7 spirals into membrane-sculpting helices. Finally, we show that ESCRT-II and ESCRT-III coassemble into ~65 nm diameter rings indicative of a cargo-sequestering supercomplex. We propose that ESCRT-III has distinct architectural stages that are modulated by ESCRT-II to mediate cargo capture and vesicle formation by ordered assembly.
Weill Institute for Cell and Molecular Biology, Cornell University, Ithaca, NY 14853, USA.
William Mike
WM
Henne
Nicholas J
NJ
Buchkovich
Yingying
Y
Zhao
Scott D
SD
Emr
D056827:Endosomal Sorting Complexes Required for Transport / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D057146:Multivesicular Bodies / Q000737:chemistry / Q000378:metabolism*; D009154:Mutation; D012441:Saccharomyces cerevisiae / Q000166:cytology / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D022161:Transport Vesicles / Q000378:metabolism
C524222:DID4 protein, S cerevisiae; D056827:Endosomal Sorting Complexes Required for Transport; C083890:SNF7 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; C533340:Vps24 protein, S cerevisiae