Mechanisms underlying lncRNA SAF-mediated survival and persistence of HIV-1 infected macrophages

点击翻译按钮获取中文摘要

Project Summary / Abstract Combination anti-retroviral therapy is highly effective in blocking HIV-1 replication and thereby new infections but it cannot eliminate reservoir cells infected prior to the treatment initiation. Most studies of HIV-1 persistence have focused on latently infected CD4+ T cells. However, tissue-resident macrophages which are self-renewable, long-lived myeloid cells, have recently been shown to sustain prolonged viremia in absence of T lymphocytes. The unique ability of macrophages to support HIV-1 replication without succumbing to virus- induced cell death makes them ideal for viral persistence. Understanding the role of viral and host factors that enable macrophages to evade cell death is needed to develop effective strategies for eliminating these persistently infected myeloid cells. By screening a panel of 90 long non-coding RNAs (lncRNA), we have recently discovered the novel role of the lncRNA SAF (FAS-AS1) in maintenance of cell survival of HIV-1 infected macrophages. This lncRNA is significantly up-regulated in HIV-1 infected human monocyte-derived macrophages (MDM) in vitro and lung alveolar macrophages (AM) in vivo. More importantly, siRNA-mediated inhibition of SAF leads to activation of apoptotic caspases in HIV-1 infected macrophages selectively, while leaving the virus-exposed yet uninfected bystander cells unaffected. This highly specific modulation of the lncRNA SAF results in a marked reduction in viral burden in the macrophage culture, emphasizing the therapeutic potential of targeting this lncRNA. Our central hypothesis is that active HIV-1 infection induces SAF up-regulation in macrophages to regulate cell survival/death pathway(s) that promote viral persistence. We propose an in-depth analysis of both the upstream regulators of SAF transcription and the downstream effectors for its function. Our specific aims are to: (1) define the viral and cellular regulators for induction of lncRNA SAF in HIV-1 infected macrophages; and (2) determine the molecular mechanisms of lncRNA SAF mediated protection of HIV-1 infected macrophages from cell death by identifying the DNA/RNA and/or protein targets of this lncRNA. We will use a panel of specific replication-stage-defective viruses in combination with state-of-the-art genetic and proteomic tools (DNA/RNA-Seq, RNA antisense purification and LC-MS) to achieve our goals. The results from this study will improve our basic scientific knowledge of the biology of a lncRNA in HIV-1 pathogenesis and persistence in macrophages. It will also provide additional modalities for targeting SAF by identifying the regulators and effectors of this lncRNA. Understanding the mechanisms underpinning SAF-mediated cell survival and viral persistence in HIV-1 infected macrophages will aid greatly in optimally exploiting the novel therapeutic potential of this lncRNA.