Abstract
BackgroundThe human immunodeficiency virus type 1 (HIV-1) Tat protein is a major viral transactivator required for HIV-1 replication. In the nucleus Tat greatly stimulates the synthesis of full-length transcripts from the HIV-1 promoter by causing efficient transcriptional elongation. Tat induces elongation by directly interacting with the bulge of the transactivation response (TAR) RNA, a hairpin-loop located at the 5'-end of all nascent viral transcripts, and by recruiting cellular transcriptional co-activators. In the cytoplasm, Tat is thought to act as a translational activator of HIV-1 mRNAs. Thus, Tat plays a central role in the regulation of HIV-1 gene expression both at the level of mRNA and protein synthesis. The requirement of Tat in these processes poses an essential question on how sufficient amounts of Tat can be made early on in HIV-1 infected cells to sustain its own synthesis. To address this issue we studied translation of the Tat mRNA in vitro and in human cells using recombinant monocistronic and dicistronic RNAs containing the 5' untranslated region (5'-UTR) of Tat RNA.ResultsThis study shows that the Tat mRNA can be efficiently translated both in vitro and in cells. Furthermore, our data suggest that translation initiation from the Tat mRNA probably occurs by a internal ribosome entry site (IRES) mechanism. Finally, we show that Tat protein can strongly stimulate translation from its cognate mRNA in a TAR dependent fashion.ConclusionThese results indicate that Tat mRNA translation is efficient and benefits from a feedback stimulation by the Tat protein. This translational control mechanism would ensure that minute amounts of Tat mRNA are sufficient to generate enough Tat protein required to stimulate HIV-1 replication.
Highlights
The human immunodeficiency virus type 1 (HIV-1) Tat protein is a major viral transactivator required for HIV-1 replication
Results reported in figure 3B show that translation in the rabbit reticulocyte lysates (RRL) of the mono 5'Glob-RNA was 5' cap-dependent, while that of the mono EMCV RNA was not
Β-galactosidase was synthesized in the context of the bicistronic Bi-EMCV RNA but clearly not synthesized when the Bi-Glob RNA was used as template
Summary
The human immunodeficiency virus type 1 (HIV-1) Tat protein is a major viral transactivator required for HIV-1 replication. The requirement of Tat in these processes poses an essential question on how sufficient amounts of Tat can be made early on in HIV-1 infected cells to sustain its own synthesis. The requirement of Tat in these processes poses an essential question on how sufficient amounts of this viral protein can be made early on in HIV-1 infected cells to sustain its own synthesis. Soon after completion of viral DNA synthesis by reverse transcriptase and before its integration into the host genome, the viral DNA can be transcribed, but this generates only low levels of fully spliced viral mRNAs encoding Tat and Nef [15] These observations led us to hypothesize that Tat mRNA is translated even under conditions where it is present in minute quantities together with a high concentration of cellular mRNAs
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