Suboptimal Provirus Expression Explains Apparent Nonrandom Cell Coinfection with HIV-1

Abstract

Despite the ability of primate lentiviruses to prevent infected cells from being reinfected, cell coinfection has occurred in the past and has shaped virus evolution by promoting the biogenesis of heterozygous virions and recombination during reverse transcription.In vitroexperiments have shown that cell coinfection with HIV is more frequent than would be expected if coinfection were a random process. A possible explanation for this bias is the heterogeneity of target cells and the preferred infection of a subpopulation. To address this question, we compared the frequency of double-positive cells measured following coincubation with green fluorescent protein (GFP) and DsRed HIV reporter viruses with that of stochastic coinfection calculated as the product of the frequencies of GFP- and DsRed-positive cells upon incubation with either reporter virus. Coinfection was more frequent than would be expected on the grounds of stochastic infection, due to the underestimation of single-infection frequencies, which mathematically decreased the calculated frequency. Indeed, when cells were incubated with either reporter virus, a fraction of the cells were scored as uninfected yet harbored a silent provirus that was reactivated upon coinfection through cross talk between viral elements. When such cross talk was avoided, experimental and calculated coinfection frequencies matched, indicating random coinfection. The proportion of infected cells harboring a silent provirus was estimated from coinfection experiments and was shown to be cell type dependent but independent of the virus entry route.