Early cytoplasmic uncoating is associated with infectivity of HIV-1

Abstract

After fusion, HIV delivers its conical capsid into the cytoplasm. To release the contained reverse-transcribing viral genome, the capsid must disassemble in a process termed uncoating. Defining the kinetics, dynamics, and cellular location of uncoating of virions leading to infection has been confounded by defective, noninfectious particles and the stochastic minefield blocking access to host DNA. We used live-cell fluorescent imaging of intravirion fluid phase markers to monitor HIV-1 uncoating at the individual particle level. We find that HIV-1 uncoating of particles leading to infection is a cytoplasmic process that occurs ∼30 min postfusion. Most, but not all, of the capsid protein is rapidly shed in tissue culture and primary target cells, independent of entry pathway. Extended time-lapse imaging with less than one virion per cell allows identification of infected cells by Gag-GFP expression and directly links individual particle behavior to infectivity, providing unprecedented insights into the biology of HIV infection.

Virological characterization of HIV‐1 CA‐NTD mutants constructed in a virus‐lineage reflected manner

“…It is also necessary to define in detail how virion-Gag disassembles at the early post-entry stage. This subject is now under vigorous investigation by many researchers (26)(27)(28)(29)(30)(31). For better understanding the HIV-1 virology and for therapeutic purpose, the above issues need to be precisely elucidated.…”

OMEGA: a software tool for the management, analysis, and dissemination of intracellular trafficking data that incorporates motion type classification and quality control

“…However, many of the fundamental image data management limitations holding back "[…] the routine application of automated image analysis […] to large volumes of information generated by digital imaging" (verbatim from: Swedlow et al, 2003) are still in place even several years after the initial identification of the problem. As a case in point, the utilization of viral particle tracking to draw direct real-time correlations between alterations in viral mobility and underlying perturbations in the viral and cellular states, remains a considerable challenge even at low-throughput, and it is difficult if not impossible to scale to the systems biology level (Arhel et al, 2006;McDonald et al, 2002;Mamede et al, 2017;Mamede and Hope, 2016;Sood et al, 2017). As a result, most virology studies to date rely on biochemical and genetic analyses conducted in bulk and on the microscopic analysis of fixed cells, which fail to capture viral heterogeneity and the complexity of viral infection processes.…”

“…It is therefore often important to report changes in the signal intensity of tracked objects (Figure 4 and Supplemental Table I). For example, when tracking dual-color enveloped viral particles carrying a membrane marker, a sudden drop in intensity might indicate fusion between the viral envelope and the acidic endocytic compartment (Mamede et al, 2017;Sood et al, 2017;Padilla-Parra et al, 2013;Sood et al, 2016;Itano et al, 2018). Alternatively, when studying endocytic trafficking, changes in intensity might inform about specific cargo sorting events (Navaroli et al, 2012).…”

Resolving the Sites of HIV Entry: Dynamin Networks Hold the Key

“…Thus, there should be a layered loss of fluorescent protein at the point of fusion and HIV core uncoating. Indeed, recent studies have stringently demonstrated that only the virions that undergo this stepwise content loss actually lead to productive HIV infection [47]. Thus, the complete one step loss of the content marker may represent membrane fusion with either an open HIV core (i.e.…”