Dynamics of HIV-1 Assembly and Release

PLoS Pathogens

Hans-Georg Kräusslich¹  Barbara Müller¹  Marko Lampe¹  Christoph Bräuchle²  Sergey Ivanchenko²  Don C. Lamb²  William J. Godinez³  Roland Eils³  Karl Rohr³ 

[1] Department of Virology, Universitätsklinikum Heidelberg, Heidelberg, Germany;Physical Chemistry, Department of Chemistry and Biochemistry, Munich Center for Integrated Protein Science (CiPSM) and Center for NanoScience, Ludwig-Maximilians-Universität München, Munich, Germany;University of Heidelberg, BIOQUANT, IPMB, and German Cancer Research Center (DKFZ), Department of Bioinformatics and Functional Genomics, Biomedical Computer Vision Group, Heidelberg, Germany

关键词: Cell membranes;    HIV-1;    Virions;    HIV;    Cytosol;    Membrane proteins;    Nucleation;    Fluorescence microscopy;   

DOI  :  10.1371/journal.ppat.1000652

学科分类：生物科学（综合）

来源: Public Library of Science

【 摘 要 】

Assembly and release of human immunodeficiency virus (HIV) occur at the plasma membrane of infected cells and are driven by the Gag polyprotein. Previous studies analyzed viral morphogenesis using biochemical methods and static images, while dynamic and kinetic information has been lacking until very recently. Using a combination of wide-field and total internal reflection fluorescence microscopy, we have investigated the assembly and release of fluorescently labeled HIV-1 at the plasma membrane of living cells with high time resolution. Gag assembled into discrete clusters corresponding to single virions. Formation of multiple particles from the same site was rarely observed. Using a photoconvertible fluorescent protein fused to Gag, we determined that assembly was nucleated preferentially by Gag molecules that had recently attached to the plasma membrane or arrived directly from the cytosol. Both membrane-bound and cytosol derived Gag polyproteins contributed to the growing bud. After their initial appearance, assembly sites accumulated at the plasma membrane of individual cells over 1–2 hours. Assembly kinetics were rapid: the number of Gag molecules at a budding site increased, following a saturating exponential with a rate constant of ∼5×10−3 s−1, corresponding to 8–9 min for 90% completion of assembly for a single virion. Release of extracellular particles was observed at ∼1,500±700 s after the onset of assembly. The ability of the virus to recruit components of the cellular ESCRT machinery or to undergo proteolytic maturation, or the absence of Vpu did not significantly alter the assembly kinetics.

【 授权许可】

CC BY