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.
Viruses. 2020 Sep 16;12(9):1032. doi: 10.3390/v12091032.
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.
内体分选复合物运输所需（ESCRT）蛋白在萌芽的细胞膜上组装，并催化它们的分裂。通过对从细胞中出芽的 HIV 病毒的实时成像，我们跟踪了 ESCRT 蛋白 ALIX、CHMP4B 和 VPS4 的募集。我们报告说，ESCRT 蛋白在病毒体组装完成后，会与病毒体短暂地共定位，持续时间为 45 ± 30 秒。我们表明，突变 Gag 的 YP 结构域（ALIX 的主要结合位点）或从细胞中耗尽 ALIX，会导致完整的 ESCRT 机制在同一个病毒体上多次募集（称为“口吃”，其中同一个病毒体上的募集次数>3）。口吃募集之间的时间间隔约为 4 ± 3 分钟，并且与野生型相互作用中单次募集的 ESCRTs 相同，停留时间（45 ± 30 秒）相同。我们的观察结果表明，ALIX 在分裂过程中发挥作用，并对 ESCRT 募集的线性模型提出质疑，这表明存在更复杂的共组装模型。
