Electrostatic lateral interactions drive ESCRT-III heteropolymer assembly

Abstract

Self-assembly of ESCRT-III complex is a critical step in all ESCRT-dependent events. ESCRT-III hetero-polymers adopt variable architectures, but the mechanisms of inter-subunit recognition in these hetero-polymers to create flexible architectures remain unclear. We demonstrate in vivo and in vitro that the Saccharomyces cerevisiae ESCRT-III subunit Snf7 uses a conserved acidic helix to recruit its partner Vps24. Charge-inversion mutations in this helix inhibit Snf7-Vps24 lateral interactions in the polymer, while rebalancing the charges rescues the functional defects. These data suggest that Snf7-Vps24 assembly occurs through electrostatic interactions on one surface, rather than through residue-to-residue specificity. We propose a model in which these cooperative electrostatic interactions in the polymer propagate to allow for specific inter-subunit recognition, while sliding of laterally interacting polymers enable changes in architecture at distinct stages of vesicle biogenesis. Our data suggest a mechanism by which interaction specificity and polymer flexibility can be coupled in membrane-remodeling heteropolymeric assemblies.

Mechanisms of negative membrane curvature sensing and generation by ESCRT III subunit Snf7

“…On the cylindrical surface, the neighboring rings on the spiral are stabilized by the interaction of helix 4 of one ring with the long helix of adjacent ring. The same type of electrostatic interaction has been recently confirmed in the hetero‐polymer of Snf7 with the ESCRTIII subunit Vps24 . The energetic properties of these preformed spirals are reported in Table .…”

“…This explains the experimental finding that helix 4 is necessary for helical oligomer formation. The same type of interaction and its importance for membrane tubulation has recently been highlighted in the Snf7‐Vps24 hetero polymer …”

An ESCRT-III Polymerization Sequence Drives Membrane Deformation and Fission

“…In the experiments analyzing the number of strands per filament ( Figures 1C and S1E), in comparison to data from Mierzwa et al (2017), higher concentrations and longer incubation times of Vps2-Vps24 were used to ensure comparability with the experiments including Vps2-Did2, for which high concentration and long incubation were required due to its low affinity for Snf7. High concentration and long incubation of Vps2-Vps24 result in increased filament bundling a phenomena we (data not shown) and other using different ESCRT-III filaments (Banjade et al, 2019) observed previously. However, the clear prevalence of multimer-of-2 strands per filament strongly indicates the occurrence of the lateral co-polymerization we described in Mierzwa et al (2017).…”

Membrane constriction and thinning by sequential ESCRT-III polymerization

“…Thus, interactions between the j and j+1 subunits along the IST1 strand provide the force that flexes the CHMP1B elbow and consequently constricts and elongates the filament. These sliding, lateral interactions to promote changes in filament architecture have also been observed for the yeast ESCRT-III proteins Snf7 and Vps24 ( 38 ).…”