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Open AccessArticle

Abrogating ALIX Interactions Results in Stuttering of the ESCRT Machinery

by Shilpa Gupta 1,2, Mourad Bendjennat 1,3,4 and Saveez Saffarian 1,2,4,*
Center for Cell and Genome Sciences, University of Utah, Salt Lake City, UT 84112, USA
Department of Biology, University of Utah, Salt Lake City, UT 84112, USA
Radiation Oncology Department, University of Miami, Miami, FL 33136, USA
Department of Physics and Astronomy, University of Utah, Salt Lake City, UT 84112, USA
Author to whom correspondence should be addressed.
Viruses 2020, 12(9), 1032;
Received: 11 August 2020 / Revised: 5 September 2020 / Accepted: 11 September 2020 / Published: 16 September 2020
(This article belongs to the Special Issue The 11th International Retroviral Nucleocapsid and Assembly Symposium)
Endosomal sorting complexes required for transport (ESCRT) proteins assemble on budding cellular membranes and catalyze their fission. Using live imaging of HIV virions budding from cells, we followed recruitment of ESCRT proteins ALIX, CHMP4B and VPS4. We report that the ESCRT proteins transiently co-localize with virions after completion of virion assembly for durations of 45 ± 30 s. We show that mutagenizing the YP domain of Gag which is the primary ALIX binding site or depleting ALIX from cells results in multiple recruitments of the full ESCRT machinery on the same virion (referred to as stuttering where the number of recruitments to the same virion >3). The stuttering recruitments are approximately 4 ± 3 min apart and have the same stoichiometry of ESCRTs and same residence time (45 ± 30 s) as the single recruitments in wild type interactions. Our observations suggest a role for ALIX during fission and question the linear model of ESCRT recruitment, suggesting instead a more complex co-assembly model. View Full-Text
Keywords: ESCRT; stuttering; HIV; ALIX; CHMP4B; VPS4 ESCRT; stuttering; HIV; ALIX; CHMP4B; VPS4
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Gupta, S.; Bendjennat, M.; Saffarian, S. Abrogating ALIX Interactions Results in Stuttering of the ESCRT Machinery. Viruses 2020, 12, 1032.

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