Cell-associated HIV RNA: a dynamic biomarker of viral persistence

Abstract

In most HIV-infected individuals adherent to modern antiretroviral therapy (ART), plasma viremia stays undetectable by clinical assays and therefore, additional virological markers for monitoring and predicting therapy responses and for measuring the degree of HIV persistence in patients on ART should be identified. For the above purposes, quantitation of cell-associated HIV biomarkers could provide a useful alternative to measurements of viral RNA in plasma. This review concentrates on cell-associated (CA) HIV RNA with the emphasis on its use as a virological biomarker. We discuss the significance of CA HIV RNA as a prognostic marker of disease progression in untreated patients and as an indicator of residual virus replication and the size of the dynamic viral reservoir in ART-treated patients. Potential value of this biomarker for monitoring the response to ART and to novel HIV eradication therapies is highlighted.

Figure 1

The essential steps in the life cycle of HIV-1. The first step is the attachment of the virus particle to receptors on the cell surface. The HIV-1 RNA genome then enters the cytoplasm as part of a nucleoprotein complex. The viral RNA genome is reverse-transcribed into a DNA duplex, which has terminal duplications known as long terminal repeats (LTRs). The linear viral DNA molecule is part of the preintegration complex that enters the nucleus. In the nucleus, unintegrated viral DNA is found in both linear and circular forms. The unintegrated circular forms of viral DNA have either one or two LTRs, are byproducts of the integration process, and are found exclusively in the nucleus. The linear unintegrated viral DNA is the precursor of integrated proviral DNA, which is a stable structure that remains indefinitely in the host-cell genome and serves as a template for viral transcription. Transcription of the proviral DNA template and alternative RNA splicing creates spliced viral RNA species encoding the viral accessory proteins, including Tat, Rev, and Nef, and the unspliced viral RNA encoding the viral structural proteins, including the Gag–Pol precursor protein. All the viral transcripts are exported into the cytoplasm, where translation and assembly and processing of the retroviral particle take place. The cycle is completed by the release of infectious retroviral particles from the cell. (Figure adapted from [12]; reproduced, with permission, from Massachusetts Medical Society © 1999).

Figure 2

Viral RNA species produced within HIV-1-infected cells and phases of HIV-1 RNA expression. (A) Viral RNA species produced within HIV-1-infected cells. HIV-1 genes are shown relative to the long terminal repeats (LTR). The viral genomic or 9-kb unspliced RNA shows the location of 5^′ (D) and 3^′ (A) splice sites. The incompletely and multiply spliced HIV-1 viral RNAs (4-kb and 2-kb size classes) are shown as black boxes. Spliced RNAs are denoted by the translated open reading frames and by the exon content. (Figure is adapted from [20]; reproduced, with permission, from American Society for Microbiology © 2008.) (B) Early and late phases of HIV-1 RNA expression. Full-length unspliced 9-kb RNA, incompletely spliced 4-kb RNA, and multiply spliced 2-kb RNA species are constitutively expressed in the nucleus. In the absence of Rev (upper panel), or when the concentration of Rev is below the threshold necessary for function, the 9-kb and 4-kb transcripts are excluded from the cytoplasm and either spliced or degraded. In contrast, the fully processed 2-kb RNA are constitutively exported to the cytoplasm and used to express Rev, Tat, and Nef. When the levels of Rev in the nucleus are sufficiently high (lower panel), the nuclear export of 9-kb and 4-kb RNAs is activated and the translation of all viral proteins ensues. (Ball and stick) The Rev response element. (Figure adapted from [21]; reproduced, with permission, from Annual Review of Microbiology © 1998).

Figure 3

Decay kinetics of cell-free and cell-associated HIV RNA upon ART initiation.