Essential N-Terminal Insertion Motif Anchors the ESCRT-III Filament during MVB Vesicle Formation

• ESCRT-III subunits mediate a topologically unique scission event
• ESCRT-III binds membrane via a basic interface and an N-terminal amphipathic helix
• The N-terminal ANCHR motif senses curvature and anchors ESCRT-III to membrane
• Vesicle formation involves maintaining an intricate balance of opposing curvatures

SummaryThe endosomal sorting complexes required for transport (ESCRTs) have emerged as key cellular machinery that drive topologically unique membrane deformation and scission. Understanding how the ESCRT-III polymer interacts with membrane, promoting and/or stabilizing membrane deformation, is an important step in elucidating this sculpting mechanism. Using a combination of genetic and biochemical approaches, both in vivo and in vitro, we identify two essential modules required for ESCRT-III-membrane association: an electrostatic cluster and an N-terminal insertion motif. Mutating either module in yeast causes cargo sorting defects in the MVB pathway. We show that the essential N-terminal insertion motif provides a stable anchor for the ESCRT-III polymer. By replacing this N-terminal motif with well-characterized membrane insertion modules, we demonstrate that the N terminus of Snf7 has been tuned to maintain the topological constraints associated with ESCRT-III-filament-mediated membrane invagination and vesicle formation. Our results provide insights into the spatially unique, ESCRT-III-mediated membrane remodeling.

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