This project focuses on HIV-1 replication in cultured cells to develop understandings of viral pathogenesis and the cellular factors that influence viral gene expression. Examples of important scientific advances that we have achieved in the 2011 to 2012 period are outlined below. RNA helicases are ubiquitous in plants and animals and function in many cellular processes. Retroviruses, such as human immunodeficiency virus (HIV-1), encode no RNA helicases in their genomes and utilize host cellular RNA helicases at various stages of their life cycle. We have continued our study of RNA helicases in HIV-1 replication. We have identified several novel RNA helicases that activate HIV-1 post-transcriptional gene expression and an RNA helicase that represses HIV-1 replication through the interferon pathway. HIV-1 Tat protein recruits host cell factors including CDK9/cyclin T1 to HIV-1 TAR RNA and thereby induces HIV-1 transcription. An interaction with host Ser/Thr protein phosphatase-1 (PP1) is critical for this function of Tat. PP1 binds to a Tat sequence, Q(35)VCF(38), which resembles the PP1-binding "RVxF" motif present on PP1-binding regulatory subunits. We have found that expression of PP1 binding peptide, a central domain of Nuclear Inhibitor of PP1, disrupted the interaction of HIV-1 Tat with PP1 and inhibited HIV-1 transcription and replication. We have identified small molecule compounds that target the "RVxF"-binding cavity of PP1 to disrupt the interaction of PP1 with Tat and inhibit HIV-1 replication. One of the compounds, 1H4, inhibited HIV-1 transcription and replication at non-cytotoxic concentrations. Our study shows that HIV- inhibition can be achieved through using small molecules to target a non-catalytic site of PP1. We have constructed several HIV-1 molecular clones, each containing a discrete cellular miRNA positioned in Nef. These retroviral genomes express the inserted miRNA and are generally replication competent in T-cells. The inserted intragenomic miRNA was observed to elicit two different consequences for HIV-1 replication. First, the expression of miRNAs with predicted target sequences in the HIV-1 genome was found to reduce viral replication. Second, in one case, where an inserted miRNA was unusually well-processed by Drosha, this processing event inhibited viral replication. Our work is the first study to examine in detail the replication competence of HIV-1 genomes that express cis-embedded miRNAs. The results indicate that a replication competent retroviral genome is not precluded from encoding and expressing a viral miRNA.

Project Start
Project End
Budget Start
Budget End
Support Year
24
Fiscal Year
2012
Total Cost
$1,136,444
Indirect Cost
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State
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Chen, Chia-Yen; Liu, Xiang; Boris-Lawrie, Kathleen et al. (2013) Cellular RNA helicases and HIV-1: insights from genome-wide, proteomic, and molecular studies. Virus Res 171:357-65
Zhang, Quan; Chen, Chia-Yen; Yedavalli, Venkat S R K et al. (2013) NEAT1 long noncoding RNA and paraspeckle bodies modulate HIV-1 posttranscriptional expression. MBio 4:e00596-12
Zhang, Quan; Jeang, Kuan-Teh (2013) Long non-coding RNAs (lncRNAs) and viral infections. Biomed Pharmacother 3:34-42
Tang, Sai-Wen; Ducroux, Aurelie; Jeang, Kuan-Teh et al. (2012) Impact of cellular autophagy on viruses: Insights from hepatitis B virus and human retroviruses. J Biomed Sci 19:92
Zheng, Yong-Hui; Jeang, Kuan-Teh; Tokunaga, Kenzo (2012) Host restriction factors in retroviral infection: promises in virus-host interaction. Retrovirology 9:112
Ammosova, Tatiana; Platonov, Maxim; Yedavalli, Venkat R K et al. (2012) Small molecules targeted to a non-catalytic "RVxF" binding site of protein phosphatase-1 inhibit HIV-1. PLoS One 7:e39481
Liu, Xiang; Houzet, Laurent; Jeang, Kuan-Teh (2012) Tombusvirus P19 RNA silencing suppressor (RSS) activity in mammalian cells correlates with charged amino acids that contribute to direct RNA-binding. Cell Biosci 2:41
Jerebtsova, Marina; Kumari, Namita; Xu, Min et al. (2012) HIV-1 Resistant CDK2-Knockdown Macrophage-Like Cells Generated from 293T Cell-Derived Human Induced Pluripotent Stem Cells. Biology (Basel) 1:175-195
Bachu, Mahesh; Yalla, Swarupa; Asokan, Mangaiarkarasi et al. (2012) Multiple NF-ýýB sites in HIV-1 subtype C long terminal repeat confer superior magnitude of transcription and thereby the enhanced viral predominance. J Biol Chem 287:44714-35
Klase, Zachary; Houzet, Laurent; Jeang, Kuan-Teh (2012) MicroRNAs and HIV-1: complex interactions. J Biol Chem 287:40884-90

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