Direct evidence for translational regulation by leader RNA and Tat protein of human immunodeficiency virus type 1.

Sengupta, Dibyendu N.; Berkhout, Ben; Gatignol, Anne; Zhou, Aimin; Silverman, Robert H.

doi:10.1073/pnas.87.19.7492

Cited by 89 publications

(74 citation statements)

References 36 publications

(61 reference statements)

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“…These data reinforce the role of the TAR element as a translational inhibitor and point out the importance of the dsRNA structure of transcripts in the efficiency of translation. It has been previously reported that the secondary structure of HIV-1 TAR affects translation efficiency by preventing the accessibility of the cap structure (31), by activating the PKRmediated phosphorylation of the initiation factor eIF-2␣ (12,32) or by both mechanisms (33,34). Our results indicate that the TAR secondary structure is a significant impediment to translational initiation.…”

Section: Discussionsupporting

confidence: 57%

The TAR RNA-binding Protein, TRBP, Stimulates the Expression of TAR-containing RNAs in Vitro and in VivoIndependently of Its Ability to Inhibit the dsRNA-dependent Kinase PKR

Dorin¹,

Bonnet²,

Bannwarth³

et al. 2003

Journal of Biological Chemistry

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TRBP (HIV-1 transactivating response (TAR) RNAbinding protein) and PKR, the interferon-induced dsRNA-regulated protein kinase, contain two dsRNA binding domains. They both bind to HIV-1 TAR RNAs through different sites. Binding to dsRNA activates PKR that phosphorylates the eukaryotic initiation factor eIF-2␣ leading to protein synthesis inhibition. TRBP and PKR can heterodimerize, which inhibits the kinase function of PKR and has a positive effect on HIV-1 expression. In this study, an in vitro reticulocyte assay revealed the poor expression of TAR containing CAT RNAs compared with CAT RNAs. Addition of TRBP restored translation efficiency of TAR-CAT RNA and decreased the phosphorylation status of eIF-2␣, confirming its role as a PKR inhibitor. Unexpectedly, eIF-2␣ was phosphorylated in the presence of TAR-CAT as well as CAT RNA devoid of the TAR structure. TRBP inhibited eIF-2␣ phosphorylation in both cases, suggesting that it restores the translation of TAR-CAT RNA independently and in addition to its ability to inhibit PKR. TRBP activity on gene expression was then analyzed in a PKRfree environment using PKR-deficient murine embryo fibroblasts. In a transient reporter gene assay, TRBP stimulated the expression of a TAR-containing luciferase 3.8-fold whereas the reporter gene with mutated TAR structures or devoid of TAR was stimulated 1.5-to 2.4-fold. Overall, the activity of TRBP2 was higher when the 5-end of the mRNA was structured and was mediated independently by each dsRBD in TRBP. Increasing concentrations of TRBP showed no significant modification of the luciferase RNA levels, suggesting that TRBP stimulates translation of TAR-containing RNAs. Therefore, TRBP is an important cellular factor for efficient translation of dsRNA containing transcripts, both by inhibiting PKR and in a PKR-independent pathway.

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Section: Discussionsupporting

confidence: 57%

The TAR RNA-binding Protein, TRBP, Stimulates the Expression of TAR-containing RNAs in Vitro and in VivoIndependently of Its Ability to Inhibit the dsRNA-dependent Kinase PKR

Dorin¹,

Bonnet²,

Bannwarth³

et al. 2003

Journal of Biological Chemistry

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“…The effect of TAR on 2-5A synthetase is abolished in the presence of HIV-1 Tat protein (Schroder et el., 1990), which binds to TAR RNA stem-loop (Muller etal., 1990). Similar effects of Tat have also been reported on PKR (SenGupta et al, 1990).…”

Section: Introductionsupporting

confidence: 66%

Modulation of the Antiviral 2-5A System in Human Immunodeficiency Virus-1-Infected CEM Cells by Propentofylline

Leuck

Kuusksalu

Kelve

et al. 1996

Antivir Chem Chemother

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2′,5′-OIigoadenylates (2-5A) play an essential role in the establishment of the antiviral state of cells exposed to virus infection. However, - after an initial increase observed in some cell lines - the activity of the interferon (IFN)-inducible, 2-5A-forming 2′,5′-oligoadenylate synthetase (2-5A synthetase) strongly decreases soon after infection of cells with the human immunodeficiency virus-1 (HIV-1). In the present report, we show that in IFN-treated human T lymphoblastoid CEM cells, the decrease in 2-5A synthetase activity had already occurred at day 1 post infection (p.i.)- At days 3 and 5 p.i., the 2-5A synthetase activity in the IFN-treated infected cells amounted to only 10-12% of that in IFN-treated uninfected control cells. The decrease in 2-5A synthetase activity was accompanied by a decrease in 2-5A synthetase mRNA and protein. We found that the decrease in 2-5A synthetase activity can be retarded by addition of the cAMP phosphodiesterase inhibitor, propentofylline. At a concentration of 30-100 μM, propentofylline displayed a significant cytoprotective and antiviral effect on HIV-1-infected CEM cells.

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“…Another interpretation, which in our view is less likely, is that Tat and ElA act separately to increase the utilization of TAR CAT mRNA. While the mechanism for the effect is unknown, it could be translational, perhaps related to the interactions of TAR RNA with the protein kinase DAI (Edery et al 1989;SenGupta and Silverman 1989;Gunnery et al 1990;SenGupta et al 1990). However, post-translational mechanisms, such as concentration-dependent changes in CAT enzyme stability, also cannot be excluded.…”

Section: Cat Rna Utilization Correlates With Rna Accumulationmentioning

confidence: 99%

Synergy between HIV-1 Tat and adenovirus E1A is principally due to stabilization of transcriptional elongation.

Laspia¹,

Rice²,

Mathews³

1990

Genes & Development

107

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We studied the combined effects of Tat and general trans-activators, such as ElA and phorbol esters, on human immunodeficiency virus-1 (HIV-1) gene expression. Interaction between these two types of trans-activators may be involved in the transition from transcriptional quiesence during viral latency to active gene expression during productive infection. ElA cooperated with Tat to produce a fourfold greater increase in accumulation of full-length, cytoplasmic HIV-1-directed RNA than is expected if they were acting additively to increase RNA accumulation. Similarily, phorbol 12-myristate 13-acetate (PMA) also cooperated with Tat to elevate HIV RNA levels synergistically. Analysis of transcription rates across the HIV-1-directed transcription unit indicated, unexpectedly, that synergy between Tat and ElA could not be accounted for by increased promoter proximal transcription rates that were merely additive. However, Tat and ElA produced a greater than additive increase in transcription rates in the 3' end of the gene. These findings imply that synergy between Tat and ElA (or other general transcriptional activators) is due principally to stabilization of transcriptional elongation. Furthermore, the observation that Tat elicits only a small increase in promoter proximal transcription in the presence of ElA suggests that the magnitude of the effect of Tat on initiation is decreased when the basal level of transcription is increased. These findings underscore the importance of the ability of Tat to stabilize elongation, as well as to stimulate initiation, in an HIV-1-directed transcription unit.

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Direct evidence for translational regulation by leader RNA and Tat protein of human immunodeficiency virus type 1.

Abstract: Translational effects of the RNA leader and

Cited by 89 publications

References 36 publications

The TAR RNA-binding Protein, TRBP, Stimulates the Expression of TAR-containing RNAs in Vitro and in VivoIndependently of Its Ability to Inhibit the dsRNA-dependent Kinase PKR

The TAR RNA-binding Protein, TRBP, Stimulates the Expression of TAR-containing RNAs in Vitro and in VivoIndependently of Its Ability to Inhibit the dsRNA-dependent Kinase PKR

Modulation of the Antiviral 2-5A System in Human Immunodeficiency Virus-1-Infected CEM Cells by Propentofylline

Synergy between HIV-1 Tat and adenovirus E1A is principally due to stabilization of transcriptional elongation.

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