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, 1:46

First Online: 27 December 2004Received: 17 December 2004Accepted: 27 December 2004


BackgroundSeveral cellular positive and negative elongation factors are involved in regulating RNA polymerase II processivity during transcription elongation in human cells. In recruiting several of these regulatory factors to the 5- long terminal repeat LTR promoter during transcription elongation, HIV-1 modulates replication of its genome in a process mediated by the virus-encoded transactivator Tat. One particular cellular regulatory factor, DSIF subunit human SPT5 hSpt5, has been implicated in both positively and negatively regulating transcriptional elongation but its role in Tat transactivation in vivo and in HIV-1 replication has not been completely elucidated.

ResultsTo understand the in vivo function of hSpt5 and define its role in Tat transactivation and HIV-1 replication, we used RNA interference RNAi to specifically knockdown hSpt5 expression by degrading hSpt5 mRNA. Short-interfering RNA siRNA designed to target hSpt5 for RNAi successfully resulted in knockdown of both hSpt5 mRNA and protein levels, and did not significantly affect cell viability. In contrast to hSpt5 knockdown, siRNA-mediated silencing of human mRNA capping enzyme, a functionally important hSpt5-interacting cellular protein, was lethal and showed a significant increase in cell death over the course of the knockdown experiment. In addition, hSpt5 knockdown led to significant decreases in Tat transactivation and inhibited HIV-1 replication, indicating that hSpt5 was required for mediating Tat transactivation and HIV-1 replication.

ConclusionsThe findings presented here showed that hSpt5 is a bona fide positive regulator of Tat transactivation and HIV-1 replication in vivo. These results also suggest that hSpt5 function in transcription regulation and mRNA capping is essential for a subset of cellular and viral genes and may not be required for global gene expression.

Electronic supplementary materialThe online version of this article doi:10.1186-1742-4690-1-46 contains supplementary material, which is available to authorized users.

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Autor: Yueh-Hsin Ping - Chia-ying Chu - Hong Cao - Jean-Marc Jacque - Mario Stevenson - Tariq M Rana


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