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Th17 cell master transcription factor RORC2 regulates HIV-1 gene expression and viral outgrowth

Tomas Raul Wiche Salinas, Yuwei Zhang, Daniele Sarnello, Alexander Zhyvoloup, Laurence Raymond Marchand, Delphine Planas, Manivel Lodha, Debashree Chatterjee, Kasia Karwacz, Sally Oxenford, Jean-Pierre Routy, Heather Amrine-Madsen, Petronela Ancuta, Ariberto Fassati

Preprint posted on 28 March 2021 https://www.biorxiv.org/content/10.1101/2021.03.27.435072v1

Article now published in Proceedings of the National Academy of Sciences at http://dx.doi.org/10.1073/pnas.2105927118

Human protein RORC2 regulates HIV-1 gene expression

Selected by Joao Mello-Vieira

Background

As reported by UNAIDS*, almost 40 million people across the world were living with HIV in 2019. Since the 1990s, the development of anti-retroviral therapies means that viral proteins can be inhibited and viral load reduced to undetectable levels. These therapies greatly increase the quality of life and longevity of people infected with HIV-1. Unfortunately, infected individuals remain at risk of relapse as reservoirs of virus-infected T lymphocytes cells throughout the body cannot be eliminated by anti-retroviral therapy.

One of these reservoirs is the mucosae, such as the lining of the gut, where the main lymphocyte population are Th17 cells. Recently, the laboratory of Ariberto Fassati discovered that two compounds that inhibit HIV-1 are also know inhibitors of the Th17 cell master transcription factor RORC2 (Zhyvoloup, Melamed, et al., 2017, PLoS Pathog.).  In this current preprint, Wiche Salinas, Zhang and colleagues explore the importance of RORC2 for persistent HIV-1 infection. By testing well-characterised RORC2 inhibitors created by the pharmaceutical company GlaxoSmithKline, the authors identify RORC2 as a host co-factor for HIV-1 infection that activates HIV-1 gene expression.

* UNAIDS: the global policy coordinator for the United Nations (UN) for Acquire Immunodeficiency Syndrome (AIDS) and HIV

 

Key findings

  • Four inhibitors of the transcription factor RORC2 also inhibited infection of Jurkat cells by HIV-1 and the replication of HIV-1 viruses in primary memory T cells. Moreover, knock-down of RORC2 in Jurkat cells and primary memory T cells decreased HIV-1 replication. Conversely, overexpression of RORC2 in both Jurkat cells and HEK293T cells (the latter do not normally express this transcription factor) enhanced HIV-1 infection. This showed that human RORC2 is important for HIV-1 infection.
  • Next, the authors wondered which stage of HIV-1 infection required RORC2. RORC2 inhibition did not decrease the reverse transcription of the viral RNA, nuclear entry nor integration of viral DNA into the genome. However, if cells that were already infected with HIV-1 were treated with RORC2 inhibitors, the level of viral mRNA was decreased, suggesting that the RORC2 is important for the expression of the viral genes
  • Since RORC2 is a transcription factor, it can activate viral gene expression by directly binding to the integrated viral DNA. To detect this, the authors performed chromatin immunoprecipitation of RORC2 in HIV-1 infected Jurkat cells and observed that RORC2 does bind to integrated viral DNA. RORC2 binds specifically in a region of the viral genome that is characterized to bind other retinoic acid receptors.
  • Lastly, analysis of memory T cells from patients with HIV-1 showed that cells that express RORC2 are enriched in viral DNA. If these infected cells from patients are incubated with RORC2 inhibitors, viral outgrowth was limited, suggesting that RORC2 inhibitors can have a clinical application for the control of HIV-1.

 

Importance and significance

The location of HIV-1 infected T cell reservoirs are mostly in the mucosae, such as the gut, where the main lymphocyte population is Th17 cells. These reservoirs are maintained since people living with HIV-1 still suffer from residual immune dysfunction, such as systemic inflammation, even during anti-retroviral therapies. Importantly, residual HIV-1 transcription inside reservoir cells is driving at least in part this persistent inflammation. This constant inflammatory status allows the virus to persist, because of constant lymphocyte proliferation in response to this stimulus. In fact, the severity of this systemic inflammation corelates with the mortality, morbidity and co-morbidities that these patients suffer from.

Th17 cells are also important for the virus during the early stages of HIV-1 infection. For example, the depletion of Th17 cells from the gut leads to the leakage of bacterial and fungal products into circulation, triggering systemic inflammation. This then forces T lymphocytes to multiply as they respond to inflammation, indirectly creating more cells that HIV-1 can infect.  Further, this ensures that there is continuous leakage of pathogenic material through the now depleted mucosal surfaces.

The results published by Wiche Salinas, Zhang and colleagues show that HIV-1 uses the master transcription factor of Th17 cells to drive its own gene expression. One important aspect the authors note is that RORC2 might not drive the expression of viral genes per se. It might work by changing the chromatin status of enhancers and promoters to a poised state. Transcription would then be driven by other Th17 transcription factors, such as NF-kB, BAFT and STAT4. These results open avenues for development of new therapies as none of the current therapies in use target viral gene expression. These new therapies could be extremely useful for eliminating lingering viral reservoirs.

Tags: aids, chip, hiv, transcription factor

Posted on: 20 April 2021

doi: https://doi.org/10.1242/prelights.28448

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Author's response

Prof. Petronela Ancuta and Prof. Ariberto Fassati shared

João Mello-Vieira (JMV): Do you think that RORC2 inhibitors can be used as a form of chemoprotection from virus infection in the first place? Or would this be too difficult to implement?

Prof. Petronela Ancuta and Prof. Ariberto Fassati (PA/AF): Prevention is possible in theory and it has been tested in clinical trials with PreP. Also, microbicides containing glycerol monolaurate, which may block CCL20 mediated recruitment of CCR6+ Th17 cells into mucosal sites, showed efficacy in simian models (AT Haase, Nature, 2010). One possibility is that RORC2 inhibitors might prevent infection of Th17 cells, which are among the first cells to be targeted at transmission. However, thorough testing will be required to understand any long-term effects of the RORC2 inhibitors on the physiological functions of Th17 cells.

 

JMV: Does HIV-1 have a preference to invade preferentially Th17 cells over other T cells? Or is it just better at multiplying inside them?

PA/AF: It has been shown in macaques infected with simian immunodeficiency virus and in humans infected with HIV-1 that Th17 cells are among the very first cells to be infected in the mucosa at the time of virus transmission. Indeed, in addition to their strategic location at mucosal sites of viral entry, Th17 cells are enriched in CCR5 and other post-entry HIV permissiveness factors (e.g., NF-kB) and also lack HIV restriction mechanisms.

 

JMV: Do Th17 cells show the same importance for HIV-2 infection? Does HIV-2 also use RORC2 to regulates its gene expression?

PA/AF: We have some preliminary results that suggest the RORC2 inhibitors also reduce HIV-2 replication but more work is needed to understand how. It is possible that human and some primate lentiviruses have evolved to use RORC2 to better propagate in Th17 cells as a way to persist.

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