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Inhibition of HIV-1 fusion with small interfering RNAs targeting the chemokine coreceptor CXCR4

Gene Therapy volume 11, pages 1703–1712 (2004)Cite this article

Abstract

RNA interference (RNAi) is an evolutionarily conserved process by which plants and animals protect their genomes utilizing small, double-stranded RNAs to degrade target RNAs in a sequence-specific manner. Post-transcriptional gene silencing by these moieties can lead to degradation of both cellular and viral RNAs. It has recently been shown that double-stranded, small interfering RNAs (siRNAs) of 21–25 nucleotides can be transfected into relevant cells to target specific RNAs. This approach was utilized to inhibit human immunodeficiency virus type I (HIV-1) infection in human cells. siRNAs with homology to a motif in the mRNA that encodes for the HIV-1 chemokine coreceptor CXCR4 was utilized. Complementary studies via immunofluorescence microscopy and fluorescence-activated cell sorting demonstrated downregulation of CXCR4 from the surface of cells transfected with the specific siRNAs. As well, siRNAs without sequence homology to CXCR4 were used as controls and demonstrated no downregulation of CXCR4. siRNAs targeted to another chemokine coreceptor, APJ, showed specificity for downregulation of APJ but had no effects on CXCR4. Transfections with siRNAs targeting CXCR4 mRNA were shown to inhibit HIV-1 envelope fusion, which is relatively resistant to most viral inhibitors targeting chemokine coreceptors. The specificity of this effect was demonstrated by the inhibition of fusion by CXCR4-tropic and dual-tropic (CXCR4 and CCR5) envelope glycoproteins from HIV-1 on CXCR4+ indicator cells, but the lack of effects by siRNAs targeting CXCR4 mRNA on dual-tropic HIV-1 envelopes in CCR5+ indicator cells utilizing these fusion assays. Interestingly, siRNAs targeting CXCR4 selectively inhibited CXCR4-tropic cell-free virus infection of human cells but at only modest levels as compared to cell:cell fusion. siRNA may be a potential molecular therapeutic approach to alter a cellular cofactor critical for infection of human cells by relevant strains of HIV-1. The targeting of a cellular cofactor, rather than the HIV-1-specific mRNAs or genomic RNA, holds promise as the rapid mutational ability of the HIV-1 genome may obviate the potential clinical use of RNAi directly against this virus.

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Acknowledgements

We thank Dr B Cullen (Duke University) for kindly providing the luciferase-reported HIV-1 proviral clones and Ms Rita M Victor and Ms Brenda O Gordon for excellent secretarial assistance. The U87MG reporter cell line was obtained from the AIDS Reagent Repository (NIH), submitted by Drs Deng and D Littman. This work was supported in part by USPHS Grants NS27405, MH58526, NS44513 and NS41864 to RJP.

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  1. N Zhou and J Fang: These two authors contributed equally to this work

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  1. Division of Infectious Diseases and Environmental Medicine, Department of Medicine, The Dorrance H Hamilton Laboratories, Center for Human Virology and Biodefense, Jefferson Medical College, Thomas Jefferson University, Philadelphia, PA, USA

    N Zhou, J Fang, M Mukhtar, E Acheampong & RJ Pomerantz

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Zhou, N., Fang, J., Mukhtar, M. et al. Inhibition of HIV-1 fusion with small interfering RNAs targeting the chemokine coreceptor CXCR4. Gene Ther 11, 1703–1712 (2004). https://doi.org/10.1038/sj.gt.3302339

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