Long-term inhibition of HIV-1 replication with RNA interference against cellular co-factors.
ABSTRACT In this study we tested whether HIV-1 replication could be inhibited by stable RNAi-mediated knockdown of cellular co-factors. Cell lines capable of expressing shRNAs against 30 candidate co-factors implicated at different steps of the viral replication cycle were generated and analyzed for effects on cell viability and inhibition of HIV-1 replication. For half of these candidate co-factors we obtained knockdown cell lines that are less susceptible to virus replication. For three co-factors (ALIX, ATG16 and TRBP) the cell lines were resistant to HIV-1 replication for up to 2 months. With these cells we could test the hypothesis that HIV-1 is not able to escape from RNAi-mediated suppression of cellular co-factors, which was indeed not detected.
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ABSTRACT: MicroRNAs (miRNAs) directly and indirectly impact tumorigenesis. To perform their myriad roles, not only must precise miRNAs be generated by miRNA machinery genes but these genes such as Drosha, DGCR8, Dicer1, XPO5, TRBP, and AGO2 also have specific expression patterns, protein binding partners, and biochemical capabilities in different cancers. The published studies have demonstrated that changeable expression levels of Drosha, DGCR8, Dicer, XPO5, AGO2 and TRBP were associated with several tumors such as breast, colorectal, gastric, lung, ovarian and prostate cancer and alterations in the miRNA machinery play important roles in the carcinogenesis of these tumors. Here, we review their biological structures and functions with an eye towards understanding they could serve as cancer biomarkers.Frontiers in Oncology 05/2014; 4:1-9.
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ABSTRACT: The chemokine receptor CXCR4 is one of the major coreceptors for human immunodeficiency virus type 1 (HIV-1) and considered as an important therapeutic target. Knockdown of CXCR4 by RNA interference has emerged as a promising strategy for combating HIV-1 infection. However, there is a potential drawback to this strategy as undesired side effects may occur due to the loss of natural function of CXCR4. In this study, we developed a novel approach using a single lentiviral vector to express simultaneously CXCR4 dual-shRNAs and an shRNA-resistant CXCR4 mutant possessing the most possible natural functions of CXCR4 and reduced HIV-1 coreceptor activity. Via this approach we achieved the replacement of endogenous CXCR4 by CXCR4 mutant P191A that could compensate the functional loss of endogenous CXCR4 and significant reduction of HIV-1 replication by 59.2 %. Besides, we demonstrated that construction of recombinant lentiviral vector using 2A peptide-based strategy has significant advantages over using additional promoter-based strategy, including increase of lentivirus titer and avoidance of promoter competition. Therefore, the novel approach to block HIV-1 coreceptor CXCR4 without impairing its normal function provides a new strategy for CXCR4-targeted therapeutics for HIV-1 infection and potential universal applications to knock down a cellular protein in non-toxic manner.Molecular biotechnology. 05/2014;
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ABSTRACT: The synthesis of proteins from viral mRNA is the first step towards viral assembly. Viruses are dependent upon the cellular translation machinery to synthesize their own proteins. The synthesis of proteins from the human immunodeficiency virus (HIV) type 1 and 2 RNAs utilize several alternative mechanisms. The regulation of viral protein production requires a constant interplay between viral requirements and the cell response to viral infection. Among the antiviral cell responses, the interferon-induced RNA activated protein kinase, PKR, regulates the cellular and viral translation. During HIV-1 infection, PKR activation is highly regulated by viral and cellular factors. The cellular TAR RNA Binding Protein, TRBP, the Adenosine Deaminase acting on RNA, ADAR1, and the PKR Activator, PACT, play important roles. Recent data show that PACT changes its function from activator to inhibitor in HIV-1 infected cells. Therefore, HIV-1 has evolved to replicate in cells in which TRBP, ADAR1 and PACT prevent PKR activation to allow efficient viral protein synthesis. This proper translation will initiate the assembly of viral particles.Virus Research 07/2014; · 2.83 Impact Factor