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ESCRT-III recognition by VPS4 ATPases

Abstract

The ESCRT (endosomal sorting complex required for transport) pathway is required for terminal membrane fission events in several important biological processes, including endosomal intraluminal vesicle formation1,2, HIV budding3 and cytokinesis4. VPS4 ATPases perform a key function in this pathway by recognizing membrane-associated ESCRT-III assemblies and catalysing their disassembly5,6,7, possibly in conjunction with membrane fission. Here we show that the microtubule interacting and transport (MIT) domains of human VPS4A and VPS4B bind conserved sequence motifs located at the carboxy termini of the CHMP1–3 class of ESCRT-III proteins. Structures of VPS4A MIT–CHMP1A and VPS4B MIT–CHMP2B complexes reveal that the C-terminal CHMP motif forms an amphipathic helix that binds in a groove between the last two helices of the tetratricopeptide-like repeat (TPR) of the VPS4 MIT domain, but in the opposite orientation to that of a canonical TPR interaction. Distinct pockets in the MIT domain bind three conserved leucine residues of the CHMP motif, and mutations that inhibit these interactions block VPS4 recruitment, impair endosomal protein sorting and relieve dominant-negative VPS4 inhibition of HIV budding. Thus, our studies reveal how the VPS4 ATPases recognize their CHMP substrates to facilitate the membrane fission events required for the release of viruses, endosomal vesicles and daughter cells.

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Figure 1: VPS4 MIT–CHMP binding interactions.
Figure 2: Structural basis for VPS4 MIT recognition of CHMP1–3.
Figure 3: The Vps4 MIT Leu64Asp mutation inhibits membrane protein sorting into the vacuolar lumen.
Figure 4: Secondary MIT Leu 64 mutations decrease endosomal recruitment and inhibition of HIV-1 budding by VPS4A proteins that lack ATP-binding activity.

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Acknowledgements

We thank R. Rich, D. Myszka and S. Endicott for support; D. Winge for amino acid analysis; J. Shaw for reagents and expertise; and S. Alam for NMR expertise. W.I.S. received funding from the NIH.

Atomic coordinates for VPS4A MIT–CHMP1A180–196, free VPS4B MIT and VPS4B MIT–CHMP2B195–213 are deposited in the Protein Data Bank under accession numbers 2jq9, 2jqh and 2jqk, respectively.

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Correspondence to Wesley I. Sundquist.

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The file contains Supplementary Tables 1-2, Suppleementary Figures 1-6 with Legends, Supplementary Discussion, Supplementary Methods and additional references. (PDF 2227 kb)

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Stuchell-Brereton, M., Skalicky, J., Kieffer, C. et al. ESCRT-III recognition by VPS4 ATPases. Nature 449, 740–744 (2007). https://doi.org/10.1038/nature06172

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