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Review
. 2023 Sep 21;15(9):1969.
doi: 10.3390/v15091969.

Transcriptional Stochasticity as a Key Aspect of HIV-1 Latency

Alexia Damour  1 , Vera Slaninova  2 , Ovidiu Radulescu  3 , Edouard Bertrand  2 , Eugenia Basyuk  1
Affiliations
Review

Transcriptional Stochasticity as a Key Aspect of HIV-1 Latency

Alexia Damour et al. Viruses. .

Abstract

This review summarizes current advances in the role of transcriptional stochasticity in HIV-1 latency, which were possible in a large part due to the development of single-cell approaches. HIV-1 transcription proceeds in bursts of RNA production, which stem from the stochastic switching of the viral promoter between ON and OFF states. This switching is caused by random binding dynamics of transcription factors and nucleosomes to the viral promoter and occurs at several time scales from minutes to hours. Transcriptional bursts are mainly controlled by the core transcription factors TBP, SP1 and NF-κb, the chromatin status of the viral promoter and RNA polymerase II pausing. In particular, spontaneous variability in the promoter chromatin creates heterogeneity in the response to activators such as TNF-α, which is then amplified by the Tat feedback loop to generate high and low viral transcriptional states. This phenomenon is likely at the basis of the partial and stochastic response of latent T cells from HIV-1 patients to latency-reversing agents, which is a barrier for the development of shock-and-kill strategies of viral eradication. A detailed understanding of the transcriptional stochasticity of HIV-1 and the possibility to precisely model this phenomenon will be important assets to develop more effective therapeutic strategies.

Keywords: HIV-1 latency; chromatin; integration site; modeling; promoter; transcription; transcription factor; transcriptional burst; transcriptional noise.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Figure 4
Figure 4
Different models of HIV-1 transcription. (A) Random telegraph model; (B–E) models without Tat feedback loop, Tat is expressed in trans in (F–H)—models with active Tat feedback loop. (H) BIR—burst initiation rate; BTR—burst termination rate; PBR—polymerase binding rate; PPRR—Promoter-Pausing Release Rate. (A,C,D) Reprinted from Tantale, K.; Garcia-Oliver, E.; Robert, M.-C.; L’Hostis, A.; Yang, Y.; Tsanov, N.; Topno, R.; Gostan, T.; Kozulic-Pirher, A.; Basu-Shrivastava, M.; et al. Stochastic Pausing at Latent HIV-1 Promoters Generates Transcriptional Bursting. Nat. Commun. 2021, 12, 4503, doi:10.1038/s41467-021-24462-5 [21]. (B) Reprinted from Tantale, K.; Mueller, F.; Kozulic-Pirher, A.; Lesne, A.; Victor, J.-M.; Robert, M.-C.; Capozi, S.; Chouaib, R.; Bäcker, V.; Mateos-Langerak, J.; et al. A Single-Molecule View of Transcription Reveals Convoys of RNA Polymerases and Multi-Scale Bursting. Nat. Commun. 2016, 7, 12248, doi:10.1038/ncomms12248 [20]. (E) Reprinted from Zambrano, S.; Loffreda, A.; Carelli, E.; Stefanelli, G.; Colombo, F.; Bertrand, E.; Tacchetti, C.; Agresti, A.; Bianchi, M.E.; Molina, N.; et al. First Responders Shape a Prompt and Sharp NF-κB-Mediated Transcriptional Response to TNF-α. iScience 2020, 23, 101529, doi:10.1016/j.isci.2020.101529 [23]. (F) Reprinted from Miller-Jensen, K.; Skupsky, R.; Shah, P.S.; Arkin, A.P.; Schaffer, D.V. Genetic Selection for Context-Dependent Stochastic Phenotypes: Sp1 and TATA Mutations Increase Phenotypic Noise in HIV-1 Gene Expression. PLoS Comput. Biol. 2013, 9, e1003135, doi:10.1371/journal.pcbi.1003135 [96]. (G) Reprinted from Weinberger, L.S.; Shenk, T. An HIV Feedback Resistor: Auto-Regulatory Circuit Deactivator and Noise Buffer. PLoS Biol. 2007, 5, e9, doi:10.1371/journal.pbio.0050009 [123]. (H) Reprinted from Bullock, M.E.; Moreno-Martinez, N.; Miller-Jensen, K. A Transcriptional Cycling Model Recapitulates Chromatin-Dependent Features of Noisy Inducible Transcription. PLoS Comput. Biol. 2022, 18, e1010152, doi:10.1371/journal.pcbi.1010152 [118]. See the text for details.
Figure 1
Figure 1
Transcriptional stochasticity of HIV-1 promoter is regulated by host factors and viral transactivator Tat. Three states of the viral promoter and two stages of transcription (host- and virus-controlled) are indicated. Nucleosome-repressed state—top; promoter-proximal pause-repressed state—middle; and activated state—bottom. During the host-controlled stage, stochastic activation of the viral promoter, influenced by the depicted factors, leads to an increase in Tat level, which triggers the Tat-controlled feedback loop to fuel viral transcription. See the text for details. Created in BioRender.
Figure 2
Figure 2
Clones with HIV-1 reporter containing Tat feedback loop display phenotypic bifurcation. (A) LTR GFP IRES Tat LTR (LGIT) reporter allows HIV-1 expression noise to be studied by measuring GFP level via FACS. (B) Jurkat clones with single integration of LGIT vector obtained via FACS sorting from low- level-GFP-expressing cells (dim sort) exhibit 3 different phenotypes after 3 weeks in culture (75% OFF; 2% Bright; 23% PheB). (C) Flow histograms of LGIT PheB clones. (D) Spontaneous reactivation and shutdown of two clones infected with HIV-1 reporter with d2EGFP in place of Nef [70]. FACS analysis of the cell populations immediately following cell sorting are shown by the black lines. The same cell populations were analyzed after 7 days (green lines). The unsorted population is shown by the red lines. (A–C) Reprinted from Weinberger, L.S.; Burnett, J.C.; Toettcher, J.E.; Arkin, A.P.; Schaffer, D.V. Stochastic Gene Expression in a Lentiviral Positive-Feedback Loop: HIV-1 Tat Fluctuations Drive Phenotypic Diversity. Cell 2005, 122, 169–182 [63], pp. 171, 173. Copyright 2023, with permission from Elsevier. (D) Reprinted from Pearson, R.; Kim, Y.K.; Hokello, J.; Lassen, K.; Friedman, J.; Tyagi, M.; Karn, J. Epigenetic Silencing of Human Immunodeficiency Virus (HIV) Transcription by Formation of Restrictive Chromatin Structures at the Viral Long Terminal Repeat Drives the Progressive Entry of HIV into Latency. J. Virol. 2008, 82, 12291–12303 [70], p. 12295.
Figure 3
Figure 3
Transcriptional bursting of HIV-1. (A) HIV-1 reporter tagged with 128 MS2 (MCP-binding sites) allows nascent and mature HIV-1 RNAs to be detected in live cells through binding of MCP-GFP, which is co-expressed in the same cell. (B) Microscopic image of the cell expressing HIV-1 reporter depicted in (A). Bright spot is an active transcription site, small spots are single RNA molecules. (C) Top—promoter ON and OFF states; bottom—RNA polymerase II initiation events during the ON states, shown as orange lines, allow convoys to be formed, which are indicated. (D) Scheme of switching of HIV-1 promoter between chromatin-repressed OFF2 state and TBP-bound OFF1 state and ON state, during which the polymerase convoys are formed under control of the mediator complex. (E) Aggregated graphs show intensity of transcription site (TS) of the HIV-1 reporter during 8h long recordings under High- and No-Tat conditions, and each line represents a recording of one cell. ON states of the promoter, which correspond to one or several transcriptional bursts, are in green, and the OFF states are in red. # Movies –movie number (A,E) Reprinted from Tantale, K.; Garcia-Oliver, E.; Robert, M.-C.; L’Hostis, A.; Yang, Y.; Tsanov, N.; Topno, R.; Gostan, T.; Kozulic-Pirher, A.; Basu-Shrivastava, M.; et al. Stochastic Pausing at Latent HIV-1 Promoters Generates Transcriptional Bursting. Nat. Commun. 2021, 12, 4503, doi:10.1038/s41467-021-24462-5 [21].
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Grants and funding

A.D. was funded by ANRS fellowship (ANRS0371b); V.S. was funded by Sidaction and ANRS fellowships (2022-1-FJC-13332, ANRS0068); E.B. (Edouard Bertrand) and O.R. were funded by ANRS (ANRS0068); E.B. (Eugenia Basyuk) was funded by Sidaction (l’aide aux equipes 13316).

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