Single-Cell and Single-Cycle Analysis of HIV-1 Replication

PLoS Pathogens

Paul D. Bieniasz¹  Fengwen Zhang¹  Mowgli Holmes² 

[1] Aaron Diamond AIDS Research Center and Laboratory of Retrovirology, The Rockefeller University, New York, New York, United States of America;Columbia University, New York, New York, United States of America

关键词: HIV-1;    Gene expression;    Virions;    Viral replication;    Fluorescence imaging;    Viral gene expression;    Reporter genes;    T cells;   

DOI  :  10.1371/journal.ppat.1004961

学科分类：生物科学（综合）

来源: Public Library of Science

【 摘 要 】

The dynamics of the late stages of the HIV-1 life cycle are poorly documented. Viral replication dynamics are typically measured in populations of infected cells, but asynchrony that is introduced during the early steps of HIV-1 replication complicates the measurement of the progression of subsequent steps and can mask replication dynamics and their variation in individual infected cells. We established microscopy-based methods to dynamically measure HIV-1-encoded reporter gene and antiviral gene expression in individual infected cells. We coupled these measurements with conventional analyses to quantify delays in the HIV-1 replication cycle imposed by the biphasic nature of HIV-1 gene expression and by the assembly-inhibiting property of the matrix domain of Gag. We further related the dynamics of restriction factor (APOBEC3G) removal to the dynamics of HIV-1 replication in individual cells. These studies provide a timeline for key events in the HIV-1 replication cycle, and reveal that the interval between the onset of early and late HIV-1 gene expression is only ~3h, but matrix causes a ~6–12h delay in the generation of extracellular virions. Interestingly, matrix delays particle assembly to a time at which APOBEC3G has largely been removed from the cell. Thus, a need to prepare infected cells to be efficient producers of infectious HIV-1 may provide an impetus for programmed delays in HIV-1 virion genesis. Our findings also emphasize the significant heterogeneity in the length of the HIV-1 replication cycle in homogenous cell populations and suggest that a typical infected cell generates new virions for only a few hours at the end of a 48h lifespan. Therefore, small changes in the lifespan of infected cells might have a large effect on viral yield in a single cycle and the overall clinical course in infected individuals.

【 授权许可】

CC BY