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The HIV-1 virion-associated protein vpr is a coactivator of the human glucocorticoid receptor.

Kino T, Gragerov A, Kopp JB, Stauber RH, Pavlakis GN, Chrousos GP - J. Exp. Med. (1999)

Bottom Line: We report that Vpr enhances the activity of glucocorticoids in lymphoid and muscle-derived cell lines by interacting directly with the glucocorticoid receptor and general transcription factors, acting as a coactivator.A mutant Vpr molecule with disruption of this coactivator signature motif lost its ability to influence transcription of glucocorticoid-responsive genes and became a dominant-negative inhibitor of Vpr, possibly by retaining its general transcription factor-binding activities.The glucocorticoid coactivator activity of Vpr may contribute to increased tissue glucocorticoid sensitivity in the absence of hypercortisolism and to the pathogenesis of AIDS.

View Article: PubMed Central - PubMed

Affiliation: Section on Pediatric Endocrinology, Developmental Endocrinology Branch, National Institute of Child Health and Human Development, Bethesda, MD 20892, USA. kinot@cc1.nichd.nih.gov

ABSTRACT
The HIV-1 virion-associated accessory protein Vpr affects both viral replication and cellular transcription, proliferation, and differentiation. We report that Vpr enhances the activity of glucocorticoids in lymphoid and muscle-derived cell lines by interacting directly with the glucocorticoid receptor and general transcription factors, acting as a coactivator. Vpr contains the signature motif LXXLL also present in cellular nuclear receptor coactivators, such as steroid receptor coactivator 1 and p300/CREB-binding protein, which mediates their interaction with the glucocorticoid and other nuclear hormone receptors. A mutant Vpr molecule with disruption of this coactivator signature motif lost its ability to influence transcription of glucocorticoid-responsive genes and became a dominant-negative inhibitor of Vpr, possibly by retaining its general transcription factor-binding activities. The glucocorticoid coactivator activity of Vpr may contribute to increased tissue glucocorticoid sensitivity in the absence of hypercortisolism and to the pathogenesis of AIDS.

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(A) MMTV (top) and synthetic GRE-containing (bottom) promoter constructs used to demonstrate dependence of Vpr effect on the presence of GREs. (B and C) Decreasing numbers of GREs in the MMTV (B) or synthetic GRE-containing (C) promoter were associated with decreasing  Vpr glucocorticoid coactivator activity. A204 cells were transfected with different amounts of pCDNA3-VPR and pMMTV-luc, pHH-luc, pM-luc,  pGRE2-E1B-CAT, pGRE2-NF1-E1B-CAT, pNF1-E1B-CAT, or pE1B-CAT. Black and white bars, mean ± SEM values obtained in the absence or  presence of 10−7 M dexamethasone, respectively. *P < 0.001. (D) The coactivator effect of Vpr depends on the presence of functional hGR in CV-1 cells.  Cells were transfected with different amounts of pCDNA3-VPR and pMMTV-luc in the presence (white bars) or absence (black bars) of pRShGRα.  For control, pRS-erbA−1 was transfected instead of pRShGRα. Bars, mean ± SEM values obtained in the presence of 10−7 M dexamethasone. *P <  0.001. (E) Detection of the GR isoforms α and β (a) or only α (b) in A204 cells transfected with pHook™-1 and either pCDNA3 (−) or pCDNA3-VPR (+). Transfection-positive cells were collected by Capture-Tec™ beads. The GR (α and β) and the GRα were detected after blotting by using antibodies that can recognize both the GR α and β (a), and specific antibodies for the GRα (b).
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Figure 2: (A) MMTV (top) and synthetic GRE-containing (bottom) promoter constructs used to demonstrate dependence of Vpr effect on the presence of GREs. (B and C) Decreasing numbers of GREs in the MMTV (B) or synthetic GRE-containing (C) promoter were associated with decreasing Vpr glucocorticoid coactivator activity. A204 cells were transfected with different amounts of pCDNA3-VPR and pMMTV-luc, pHH-luc, pM-luc, pGRE2-E1B-CAT, pGRE2-NF1-E1B-CAT, pNF1-E1B-CAT, or pE1B-CAT. Black and white bars, mean ± SEM values obtained in the absence or presence of 10−7 M dexamethasone, respectively. *P < 0.001. (D) The coactivator effect of Vpr depends on the presence of functional hGR in CV-1 cells. Cells were transfected with different amounts of pCDNA3-VPR and pMMTV-luc in the presence (white bars) or absence (black bars) of pRShGRα. For control, pRS-erbA−1 was transfected instead of pRShGRα. Bars, mean ± SEM values obtained in the presence of 10−7 M dexamethasone. *P < 0.001. (E) Detection of the GR isoforms α and β (a) or only α (b) in A204 cells transfected with pHook™-1 and either pCDNA3 (−) or pCDNA3-VPR (+). Transfection-positive cells were collected by Capture-Tec™ beads. The GR (α and β) and the GRα were detected after blotting by using antibodies that can recognize both the GR α and β (a), and specific antibodies for the GRα (b).

Mentions: To show the dependency of the Vpr effect on the presence of GREs, we used three MMTV deletion mutants, four GRE-containing promoter constructs, the simian virus 40 (SV40) and Rous sarcoma virus (RSV) promoters, and the human β-actin promoter, which contain no recognizable GREs. Because the NF1 site is important for the full activation of the MMTV promoter, we used promoter constructs containing a synthetic NF1 site (Fig. 2 A). As shown in Fig. 2, B and C, the coactivator effect of Vpr depended on the presence of GREs. Decreasing the numbers of GREs in the MMTV or in synthetic GRE promoters was associated with diminishing Vpr coactivator activity. Vpr had no or minimal effect on the synthetic promoters not containing GRE sites and on the SV40, RSV, and β-actin promoters (data not shown). We also used CV-1 cells, which contain no functional GR, to show the requirement of the GR for the coactivator effect of Vpr on the MMTV promoter. Vpr-dependent activation could be observed only when CV-1 cells were cotransfected with the GRα expression vector pRShGRα (Fig. 2 D).


The HIV-1 virion-associated protein vpr is a coactivator of the human glucocorticoid receptor.

Kino T, Gragerov A, Kopp JB, Stauber RH, Pavlakis GN, Chrousos GP - J. Exp. Med. (1999)

(A) MMTV (top) and synthetic GRE-containing (bottom) promoter constructs used to demonstrate dependence of Vpr effect on the presence of GREs. (B and C) Decreasing numbers of GREs in the MMTV (B) or synthetic GRE-containing (C) promoter were associated with decreasing  Vpr glucocorticoid coactivator activity. A204 cells were transfected with different amounts of pCDNA3-VPR and pMMTV-luc, pHH-luc, pM-luc,  pGRE2-E1B-CAT, pGRE2-NF1-E1B-CAT, pNF1-E1B-CAT, or pE1B-CAT. Black and white bars, mean ± SEM values obtained in the absence or  presence of 10−7 M dexamethasone, respectively. *P < 0.001. (D) The coactivator effect of Vpr depends on the presence of functional hGR in CV-1 cells.  Cells were transfected with different amounts of pCDNA3-VPR and pMMTV-luc in the presence (white bars) or absence (black bars) of pRShGRα.  For control, pRS-erbA−1 was transfected instead of pRShGRα. Bars, mean ± SEM values obtained in the presence of 10−7 M dexamethasone. *P <  0.001. (E) Detection of the GR isoforms α and β (a) or only α (b) in A204 cells transfected with pHook™-1 and either pCDNA3 (−) or pCDNA3-VPR (+). Transfection-positive cells were collected by Capture-Tec™ beads. The GR (α and β) and the GRα were detected after blotting by using antibodies that can recognize both the GR α and β (a), and specific antibodies for the GRα (b).
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Figure 2: (A) MMTV (top) and synthetic GRE-containing (bottom) promoter constructs used to demonstrate dependence of Vpr effect on the presence of GREs. (B and C) Decreasing numbers of GREs in the MMTV (B) or synthetic GRE-containing (C) promoter were associated with decreasing Vpr glucocorticoid coactivator activity. A204 cells were transfected with different amounts of pCDNA3-VPR and pMMTV-luc, pHH-luc, pM-luc, pGRE2-E1B-CAT, pGRE2-NF1-E1B-CAT, pNF1-E1B-CAT, or pE1B-CAT. Black and white bars, mean ± SEM values obtained in the absence or presence of 10−7 M dexamethasone, respectively. *P < 0.001. (D) The coactivator effect of Vpr depends on the presence of functional hGR in CV-1 cells. Cells were transfected with different amounts of pCDNA3-VPR and pMMTV-luc in the presence (white bars) or absence (black bars) of pRShGRα. For control, pRS-erbA−1 was transfected instead of pRShGRα. Bars, mean ± SEM values obtained in the presence of 10−7 M dexamethasone. *P < 0.001. (E) Detection of the GR isoforms α and β (a) or only α (b) in A204 cells transfected with pHook™-1 and either pCDNA3 (−) or pCDNA3-VPR (+). Transfection-positive cells were collected by Capture-Tec™ beads. The GR (α and β) and the GRα were detected after blotting by using antibodies that can recognize both the GR α and β (a), and specific antibodies for the GRα (b).
Mentions: To show the dependency of the Vpr effect on the presence of GREs, we used three MMTV deletion mutants, four GRE-containing promoter constructs, the simian virus 40 (SV40) and Rous sarcoma virus (RSV) promoters, and the human β-actin promoter, which contain no recognizable GREs. Because the NF1 site is important for the full activation of the MMTV promoter, we used promoter constructs containing a synthetic NF1 site (Fig. 2 A). As shown in Fig. 2, B and C, the coactivator effect of Vpr depended on the presence of GREs. Decreasing the numbers of GREs in the MMTV or in synthetic GRE promoters was associated with diminishing Vpr coactivator activity. Vpr had no or minimal effect on the synthetic promoters not containing GRE sites and on the SV40, RSV, and β-actin promoters (data not shown). We also used CV-1 cells, which contain no functional GR, to show the requirement of the GR for the coactivator effect of Vpr on the MMTV promoter. Vpr-dependent activation could be observed only when CV-1 cells were cotransfected with the GRα expression vector pRShGRα (Fig. 2 D).

Bottom Line: We report that Vpr enhances the activity of glucocorticoids in lymphoid and muscle-derived cell lines by interacting directly with the glucocorticoid receptor and general transcription factors, acting as a coactivator.A mutant Vpr molecule with disruption of this coactivator signature motif lost its ability to influence transcription of glucocorticoid-responsive genes and became a dominant-negative inhibitor of Vpr, possibly by retaining its general transcription factor-binding activities.The glucocorticoid coactivator activity of Vpr may contribute to increased tissue glucocorticoid sensitivity in the absence of hypercortisolism and to the pathogenesis of AIDS.

View Article: PubMed Central - PubMed

Affiliation: Section on Pediatric Endocrinology, Developmental Endocrinology Branch, National Institute of Child Health and Human Development, Bethesda, MD 20892, USA. kinot@cc1.nichd.nih.gov

ABSTRACT
The HIV-1 virion-associated accessory protein Vpr affects both viral replication and cellular transcription, proliferation, and differentiation. We report that Vpr enhances the activity of glucocorticoids in lymphoid and muscle-derived cell lines by interacting directly with the glucocorticoid receptor and general transcription factors, acting as a coactivator. Vpr contains the signature motif LXXLL also present in cellular nuclear receptor coactivators, such as steroid receptor coactivator 1 and p300/CREB-binding protein, which mediates their interaction with the glucocorticoid and other nuclear hormone receptors. A mutant Vpr molecule with disruption of this coactivator signature motif lost its ability to influence transcription of glucocorticoid-responsive genes and became a dominant-negative inhibitor of Vpr, possibly by retaining its general transcription factor-binding activities. The glucocorticoid coactivator activity of Vpr may contribute to increased tissue glucocorticoid sensitivity in the absence of hypercortisolism and to the pathogenesis of AIDS.

Show MeSH
Related in: MedlinePlus