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Negative feedback inhibition of HIV-1 by TAT-inducible expression of siRNA

Abstract

Here we demonstrate that an inducible anti-HIV short hairpin RNA (shRNA) expressed from a Pol II promoter inhibits HIV-1 gene expression in mammalian cells. Our strategy is based on a promoter system in which the HIV-1 LTR is fused to the Drosophila hsp70 minimal heat shock promoter. This system is inducible by HIV-1 TAT, which functions in a negative feedback loop to activate transcription of an shRNA directed against HIV-1 rev. Upon induction the shRNA is processed to an siRNA that guides inhibition of HIV replication in cultured T-lymphocytes and hematopoietic stem cell–derived monocytes. The fusion promoter system may be safer than drug-inducible systems for shRNA-mediated gene therapy against HIV as the shRNAs are only expressed following HIV infection.

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Figure 1: Expression of shRNA from a Pol II promoter (a) shRNA sequence and expression cassettes.
Figure 2: HIV-1 TAT inducible siRNA system (a) Schematic representation of LTRhsp-shRNA.
Figure 3: Inhibition of HIV-1 replication.

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Acknowledgements

This research was supported by National Institutes of Health grants AI 29329, AI42552 and HL074704.

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Correspondence to John J Rossi.

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The authors declare no competing financial interests.

Supplementary information

Supplementary Fig. 1

Transcription occurs from both, HIV-1 LTR and mhsp70 TATA boxes. (PDF 79 kb)

Supplementary Fig. 2

Partial processing of shRNA loop. (PDF 88 kb)

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Unwalla, H., Li, MJ., Kim, J. et al. Negative feedback inhibition of HIV-1 by TAT-inducible expression of siRNA. Nat Biotechnol 22, 1573–1578 (2004). https://doi.org/10.1038/nbt1040

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  • DOI: https://doi.org/10.1038/nbt1040

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