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Diverse effects of interferon alpha on the establishment and reversal of HIV latency.

ABSTRACT: HIV latency is the major barrier to a cure for people living with HIV (PLWH) on antiretroviral therapy (ART) because the virus persists in long-lived non-proliferating and proliferating latently infected CD4+ T cells. Latently infected CD4+ T cells do not express viral proteins and are therefore not visible to immune mediated clearance. Therefore, identifying interventions that can reverse latency and also enhance immune mediated clearance is of high interest. Interferons (IFNs) have multiple immune enhancing effects and can inhibit HIV replication in activated CD4+ T cells. However, the effects of IFNs on the establishment and reversal of HIV latency is not understood. Using an in vitro model of latency, we demonstrated that plasmacytoid dendritic cells (pDC) inhibit the establishment of HIV latency through secretion of type I IFN?, IFN? and IFN? but not IFN? or type III IFN?1 and IFN?3. However, once latency was established, IFN? but no other IFNs were able to efficiently reverse latency in both an in vitro model of latency and CD4+ T cells collected from PLWH on suppressive ART. Binding of IFN? to its receptor expressed on primary CD4+ T cells did not induce activation of the canonical or non-canonical NF?B pathway but did induce phosphorylation of STAT1, 3 and 5 proteins. STAT5 has been previously demonstrated to bind to the HIV long terminal repeat and activate HIV transcription. We demonstrate diverse effects of interferons on HIV latency with type I IFN?; inhibiting the establishment of latency but also reversing HIV latency once latency is established.

SUBMITTER: Van der Sluis RM

PROVIDER: S-EPMC7065813 | biostudies-literature | 2020 Feb

REPOSITORIES: biostudies-literature

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Diverse effects of interferon alpha on the establishment and reversal of HIV latency.

Van der Sluis Renée M RM Zerbato Jennifer M JM Rhodes Jake W JW Pascoe Rachel D RD Solomon Ajantha A Kumar Nitasha A NA Dantanarayana Ashanti I AI Tennakoon Surekha S Dufloo Jérémy J McMahon James J Chang Judy J JJ Evans Vanessa A VA Hertzog Paul J PJ Jakobsen Martin R MR Harman Andrew N AN Lewin Sharon R SR Cameron Paul U PU

PLoS pathogens 20200228 2

HIV latency is the major barrier to a cure for people living with HIV (PLWH) on antiretroviral therapy (ART) because the virus persists in long-lived non-proliferating and proliferating latently infected CD4+ T cells. Latently infected CD4+ T cells do not express viral proteins and are therefore not visible to immune mediated clearance. Therefore, identifying interventions that can reverse latency and also enhance immune mediated clearance is of high interest. Interferons (IFNs) have multiple im ...[more]

PMID: 32109259

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Project description:Quantification of cell associated HIV RNA (ca-RNA) is one of the most important and commonly used methods to evaluate the performance of latency-reversing agents (LRAs). Copies of HIV RNA measured by qPCR, are often normalized to the input RNA or cell number. However, these could be affected by biological variability and/or technical errors, which can be avoided by using an internal reference gene. To obtain reliable data, it is essential to select stable reference genes (RGs) of which the expression is not influenced by biological variability, the type of cells, or the LRAs used. However, to date, no study has carefully evaluated RG stability following LRA exposure. We analyzed the stability of six widely used RGs (GAPDH, TBP, YWHAZ, UBE2D2, HPRT1 and RPL27A) in human PBMC and CD4+ T cells. LRA exposure significantly influenced the stability of these RGs. Overall, TBP, UBE2D2, and RPL27A were the most stable RGs in all tested conditions. TBP was generally the most stable RG whereas GAPDH varied the most. Finally, we evaluated the impact of applying different RG normalizers to host genes and HIV ca-RNA data. Altered results were observed both in host and HIV gene expression when unstable RGs were used. Our data underline the importance of testing the stability of RGs utilized to evaluate LRA-induced HIV ca-RNA expression. To our knowledge, this is the first careful evaluation of the stability of RGs after LRA exposure and will significantly contribute to the quality of data analysis in regard to gene expression.IMPORTANCELatency-reversing agents (LRAs) are ubiquitously used in the "shock-and-kill" HIV cure strategy and their performance is often evaluated by ex-vivo quantification of cell associated HIV RNA. HIV RNA, measured by qPCR, is often normalized to internal reference genes, but the expression of these genes should not be influenced by the experimental settings. We found that treatment of human PBMC and CD4+ T cells with LRAs significantly altered the expression of several commonly used reference genes, such as GAPDH. Finally, we evaluate the impact of different reference genes on normalization of host genes and HIV cell associated RNA expression and demonstrated that using unstable reference genes dramatically altered experimental outcome. Our data highlight the importance of using reference genes that are unaffected by LRAs under study to correctly evaluate host gene and cell associated HIV RNA expression induced by latency-reversing agents.

Dataset Information

Diverse effects of interferon alpha on the establishment and reversal of HIV latency.

Publications

Diverse effects of interferon alpha on the establishment and reversal of HIV latency.

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