1887

Abstract

Transcriptional regulation of the human immunodeficiency virus type 1 (HIV-1) is a complex event that requires the cooperative action of both viral (e.g. Tat) and cellular (e.g. C/EBP, NF-B) factors. The HIV-1 Tat protein recruits the human positive transcription elongation factor P-TEFb, consisting of cdk9 and cyclin T1, to the HIV-1 transactivation response (TAR) region. In the absence of TAR, Tat activates the HIV-1 long terminal repeat (LTR) through its association with several cellular factors including C/EBP. C/EBP is a member of the CCAAT/enhancer-binding protein family of transcription factors and has been shown to be a critical transcriptional regulator of HIV-1 LTR. We examined whether Tat–C/EBP association requires the presence of the P-TEFb complex. Using immunoprecipitation followed by Western blot, we demonstrated that C/EBP–cyclin T1 association requires the presence of cdk9. Further, due to its instability, cdk9 was unable to physically interact with C/EBP in the absence of cyclin T1 or Tat. Using kinase assays, we demonstrated that cdk9, but not a cdk9 dominant-negative mutant (cdk9-dn), phosphorylates C/EBP. Our functional data show that co-transfection of C/EBP and cdk9 leads to an increase in HIV-1 gene expression when compared to C/EBP alone. Addition of C/EBP homologous protein (CHOP) inhibits C/EBP transcriptional activity in the presence and absence of cdk9 and causes a delay in HIV-1 replication in T-cells. Together, our data suggest that Tat–C/EBP association is mediated through cdk9, and that phosphorylated C/EBP may influence AIDS progression by increasing expression of HIV-1 genes.

Loading

Article metrics loading...

/content/journal/jgv/10.1099/vir.0.82487-0
2007-02-01
2021-05-12
Loading full text...

Full text loading...

/deliver/fulltext/jgv/88/2/631.html?itemId=/content/journal/jgv/10.1099/vir.0.82487-0&mimeType=html&fmt=ahah

References

  1. Abraham S., Sweet T., Sawaya B. E., Rappaport J., Khalili K., Amini S. 2005; Cooperative interaction of C/EBP beta and Tat modulates MCP-1 gene transcription in astrocytes. J Neuroimmunol 160:219–227 [CrossRef]
    [Google Scholar]
  2. Akira S., Isshiki H., Sugita T., Tanabe O., Kinoshita S., Nishio Y., Nakajima T., Hirano T., Kishimoto T. 1990; A nuclear factor for IL-6 expression (NF-IL6) is a member of a C/EBP family. EMBO J 9:1897–1906
    [Google Scholar]
  3. Amini S., Clavo A., Nadraga Y., Giordano A., Khalili K., Sawaya B. E. 2002; Interplay between cdk9 and NF-kappaB factors determines the level of HIV-1 gene transcription in astrocytic cells. Oncogene 21:5797–5803 [CrossRef]
    [Google Scholar]
  4. Amini S., Saunders M., Kelley K., Khalili K., Sawaya B. E. 2004; Interplay between HIV-1 Vpr and Sp1 modulates p21(WAF1) gene expression in human astrocytes. J Biol Chem 279:46046–46056 [CrossRef]
    [Google Scholar]
  5. An M. R., Hsieh C. C., Reisner P. D., Rabek J. P., Scott S. G., Kuninger D. T., Papaconstantinou J. 1996; Evidence for posttranscriptional regulation of C/EBPalpha and C/EBPbeta isoform expression during the lipopolysaccharide-mediated acute-phase response. Mol Cell Biol 16:2295–2306
    [Google Scholar]
  6. Ayer D. E., Kretzner L., Eisenman R. N. 1993; Mad: a heterodimeric partner for Max that antagonizes Myc transcriptional activity. Cell 72:211–222 [CrossRef]
    [Google Scholar]
  7. Blackwood E. M., Eisenman N. R. 1991; Max: a helix-loop-helix zipper protein that forms a sequence-specific DNA-binding complex with Myc. Science 251:1211–1217 [CrossRef]
    [Google Scholar]
  8. Brady J., Kashanchi F. 2005; Tat gets the ‘green’ light on transcription initiation. Retrovirology 2:69 [CrossRef]
    [Google Scholar]
  9. Buck M., Poli V., Hunter T., Chojkier M. 2001; C/EBPbeta phosphorylation by RSK creates a functional XEXD caspase inhibitory box critical for cell survival. Mol Cell 8:807–816 [CrossRef]
    [Google Scholar]
  10. Cao Z., Umek R. M., McKnight S. L. 1991; Regulated expression of three C/EBP isoforms during adipose conversion of 3T3-L1 cells. Genes Dev 5:1538–1552 [CrossRef]
    [Google Scholar]
  11. Chen R., Yang Z., Zhou Q. 2004; Phosphorylated positive transcription elongation factor b (P-TEFb) is tagged for inhibition through association with 7SK snRNA. J Biol Chem 279:4153–4160
    [Google Scholar]
  12. Claudio P. P., Cui J., Ghafouri M., Mariano C., White M. K., Safak M., Sheffield J. B., Giordano A., Khalili K. other authors 2006; Cdk9 phosphorylates p53 on serine 392 independently of CKII. J Cell Physiol 208:602–612 [CrossRef]
    [Google Scholar]
  13. Coyle-Rink J., Sweet T., Abraham S., Sawaya B., Batuman O., Khalili K., Amini S. 2002; Interaction between TGFbeta signaling proteins and C/EBP controls basal and Tat-mediated transcription of HIV-1 LTR in astrocytes. Virology 299:240–247 [CrossRef]
    [Google Scholar]
  14. Dang C. V., Dolde C., Gillison M. C., Kato G. J. 1992; Discrimination between related DNA sites by a single amino acid residue of Myc-related basic-helix-loop-helix proteins. Proc Natl Acad Sci U S A 89:599–602 [CrossRef]
    [Google Scholar]
  15. De Falco G., Bagella L., Claudio P. P., De Luca A., Fu Y., Calabretta B., Sala A., Giordano A. 2000; Physical interaction between CDK9 and B-Myb results in suppression of B-Myb gene autoregulation. Oncogene 19:373–379 [CrossRef]
    [Google Scholar]
  16. Descombes P., Schibler U. 1991; A liver-enriched transcriptional activator protein, LAP, and transcriptional inhibitory protein, LIP, are translated from the same mRNA. Cell 67:569–579 [CrossRef]
    [Google Scholar]
  17. Falco G. D., Neri L. M., Falco M. D., Bellan C., Yu Z., Luca A. D., Leoncini L., Giordano A. 2002; Cdk9, a member of the cdc2-like family of kinases, binds to gp130, the receptor of the IL-6 family of cytokines. Oncogene 21:7464–7470 [CrossRef]
    [Google Scholar]
  18. Fraldi A., Varrone F., Napolitano G., Michels A. A., Majello B., Bensaude O., Lania L. 2005; Inhibition of Tat activity by the HEXIM1 protein. Retrovirology 2:42 [CrossRef]
    [Google Scholar]
  19. Gao H., Parkin S., Johnson P. F., Schwartz R. C. 2002; C/EBP gamma has a stimulatory role on the IL-6 and IL-8 promoters. J Biol Chem 277:38827–38837 [CrossRef]
    [Google Scholar]
  20. Garber M. E., Wei P., KewalRamani V. N., Mayall T. P., Herrmann C. H., Rice A. P., Littman D. R., Jones K. A. 1998; The interaction between HIV-1 Tat and human cyclin T1 requires zinc and a critical cysteine residue that is not conserved in the murine CycT1 protein. Genes Dev 12:3512–3527 [CrossRef]
    [Google Scholar]
  21. Garriga J., Graña X. 2004; Cellular control of gene expression by T-type cyclin/CDK9 complexes. Gene 337:15–23 [CrossRef]
    [Google Scholar]
  22. Ghafouri M., Amini S., Khalili K., Sawaya B. E. 2006; HIV-1 associated dementia: symptoms and causes. Retrovirology 3:28 [CrossRef]
    [Google Scholar]
  23. Graña X., De Luca A., Sang N., Fu Y., Claudio P. P., Rosenblatt J., Morgan D. O., Giordano A. 1994; PITALRE, a nuclear CDC2-related protein kinase that phosphorylates the retinoblastoma protein in vitro. Proc Natl Acad Sci U S A 91:3834–3838 [CrossRef]
    [Google Scholar]
  24. Henderson A. J., Calame K. L. 1997; CCAAT/enhancer binding protein (C/EBP) sites are required for HIV-1 replication in primary macrophages but not CD4(+) T cells. Proc Natl Acad Sci U S A 94:8714–8719 [CrossRef]
    [Google Scholar]
  25. Henderson A. J., Zou X., Calame K. L. 1995; C/EBP proteins activate transcription from the human immunodeficiency virus type 1 long terminal repeat in macrophages/monocytes. J Virol 69:5337–5344
    [Google Scholar]
  26. Hogan T. H., Nonnemacher M. R., Krebs F. C., Henderson A., Wigdahl B. 2003; HIV-1 Vpr binding to HIV-1 LTR C/EBP cis -acting elements and adjacent regions is sequence-specific. Biomed Pharmacother 57:41–48 [CrossRef]
    [Google Scholar]
  27. Hohaus S., Petrovick M. S., Voso M. T., Sun Z., Zhang D. E., Tenen D. G. 1995; PU.1 (Spi-1) and C/EBP alpha regulate expression of the granulocyte-macrophage colony-stimulating factor receptor alpha gene. Mol Cell Biol 15:5830–5845
    [Google Scholar]
  28. Jeffrey P. D., Russo A. A., Polyak K., Gibbs E., Hurwitz J., Massague J., Pavletich N. P. 1995; Mechanism of CDK activation revealed by the structure of a cyclinA-CDK2 complex. Nature 376:313–320 [CrossRef]
    [Google Scholar]
  29. Kowenz-Leutz E., Twamley G., Ansieau S., Leutz A. 1994; Novel mechanism of C/EBP beta (NF-M) transcriptional control: activation through derepression. Genes Dev 8:2781–2791 [CrossRef]
    [Google Scholar]
  30. Lee E. S., Zhou H., Henderson A. J. 2001; Endothelial cells enhance human immunodeficiency virus type 1 replication in macrophages through a C/EBP-dependent mechanism. J Virol 75:9703–9712 [CrossRef]
    [Google Scholar]
  31. Lekstrom-Himes J., Xanthopoulos K. G. 1998; Biological role of the CCAAT/enhancer-binding protein family of transcription factors. J Biol Chem 273:28545–28548 [CrossRef]
    [Google Scholar]
  32. Liou L. Y., Herrmann C. H., Rice A. P. 2004; Human immunodeficiency virus type 1 infection induces cyclin T1 expression in macrophages. J Virol 78:8114–8119 [CrossRef]
    [Google Scholar]
  33. MacLachlan T. K., Sang N., De Luca A., Puri P. L., Levrero M., Giordano A. 1998; Binding of CDK9 to TRAF2. J Cell Biochem 71:467–478 [CrossRef]
    [Google Scholar]
  34. Marshall R. M., Graña X. 2006; Mechanisms controlling CDK9 activity. Front Biosci 11:2598–2613 [CrossRef]
    [Google Scholar]
  35. Michael N. L., D'Arcy L., Ehrenberg P. K., Redfield R. R. 1994; Naturally occurring genotypes of the human immunodeficiency virus type 1 long terminal repeat display a wide range of basal and Tat-induced transcriptional activities. J Virol 68:3163–3174
    [Google Scholar]
  36. Michels A. A., Fraldi A., Li Q., Adamson T. E., Bonnet F., Nguyen V. T., Sedore S. C., Price J. P., Price D. H. other authors 2004; Binding of the 7SK snRNA turns the HEXIM1 protein into a P-TEFb (CDK9/cyclin T) inhibitor. EMBO J 23:2608–2619 [CrossRef]
    [Google Scholar]
  37. Murre C., McCaw P. S., Vaessin H., Caudy M., Jan L. Y., Jan Y. N., Cabrera C. V., Buskin J. N., Hauschka S. D. other authors 1989; Interactions between heterologous helix-loop-helix proteins generate complexes that bind specifically to a common DNA sequence. Cell 58:537–544 [CrossRef]
    [Google Scholar]
  38. Nakajima T., Kinoshita S., Sasagawa T., Sasaki K., Naruto M., Kishimoto T., Akira S. 1993; Phosphorylation at threonine-235 by a ras -dependent mitogen-activated protein kinase cascade is essential for transcription factor NF-IL6. Proc Natl Acad Sci U S A 90:2207–2211 [CrossRef]
    [Google Scholar]
  39. Nerlov C., Ziff E. B. 1995; CCAAT/enhancer binding protein-alpha amino acid motifs with dual TBP and TFIIB binding ability co-operate to activate transcription in both yeast and mammalian cells. EMBO J 14:4318–4328
    [Google Scholar]
  40. Nguyen V. T., Kiss T., Michels A. A., Bensaude O. 2001; 7SK small nuclear RNA binds to and inhibits the activity of CDK9/cyclin T complexes. Nature 414:322–325 [CrossRef]
    [Google Scholar]
  41. Pei Y., Schwer B., Shuman S. 2003; Interactions between fission yeast Cdk9, its cyclin partner Pch1, and mRNA capping enzyme Pct1 suggest an elongation checkpoint for mRNA quality control. J Biol Chem 278:7180–7188 [CrossRef]
    [Google Scholar]
  42. Peruzzi F. 2006; The multiple functions of HIV-1 Tat: proliferation versus apoptosis. Front Biosci 11:708–717 [CrossRef]
    [Google Scholar]
  43. Peterlin B. M., Price D. H. 2006; Controlling the elongation phase of transcription with P-TEFb. Mol Cell 23:297–305 [CrossRef]
    [Google Scholar]
  44. Pines J. 1994; The cell cycle kinases. Semin Cancer Biol 5:305–313
    [Google Scholar]
  45. Poli V., Mancini F. P., Cortese R. 1990; IL-6DBP, a nuclear protein involved in interleukin-6 signal transduction, defines a new family of leucine zipper proteins related to C/EBP. Cell 63:643–653 [CrossRef]
    [Google Scholar]
  46. Pope R. M., Leutz A., Ness S. A. 1994; C/EBP beta regulation of the tumor necrosis factor alpha gene. J Clin Invest 94:1449–1455 [CrossRef]
    [Google Scholar]
  47. Ramji D. P., Foka P. 2002; CCAAT/enhancer-binding proteins: structure, function and regulation. Biochem J 365:561–575
    [Google Scholar]
  48. Roman C., Platero J. S., Shuman J., Calame K. 1990; Ig/EBP-1: a ubiquitously expressed immunoglobulin enhancer binding protein that is similar to C/EBP and heterodimerizes with C/EBP. Genes Dev 4:1404–1415 [CrossRef]
    [Google Scholar]
  49. Ron D., Habener J. F. 1992; CHOP, a novel developmentally regulated nuclear protein that dimerizes with transcription factors C/EBP and LAP and functions as a dominant-negative inhibitor of gene transcription. Genes Dev 6:439–453 [CrossRef]
    [Google Scholar]
  50. Rossi A., Mukerjee R., Ferrante P., Khalili K., Amini S., Sawaya B. E. 2006; Human immunodeficiency virus type 1 Tat prevents dephosphorylation of Sp1 by TCF-4 in astrocytes. J Gen Virol 87:1613–1623 [CrossRef]
    [Google Scholar]
  51. Ruocco M. R., Chen X., Ambrosino C., Dragonetti E., Liu W., Mallardo M., De Falco G., Palmieri C., Franzoso G. other authors 1996; Regulation of HIV-1 long terminal repeats by interaction of C/EBP(NF-IL6) and NF-kappaB/Rel transcription factors. J Biol Chem 271:22479–22486 [CrossRef]
    [Google Scholar]
  52. Sawaya B. E., Khalili K., Gordon J., Taube R., Amini S. 2000; Cooperative interaction between HIV-1 regulatory proteins Tat and Vpr modulates transcription of the viral genome. J Biol Chem 275:35209–35214 [CrossRef]
    [Google Scholar]
  53. Sebastian T., Johnson P. F. 2006; Stop and go: anti-proliferative and mitogenic functions of the transcription factor C/EBPbeta. Cell Cycle 5:953–957 [CrossRef]
    [Google Scholar]
  54. Sekine O., Nishio Y., Egawa K., Nakamura T., Maegawa H., Kashiwagi A. 2002; Insulin activates CCAAT/enhancer binding proteins and proinflammatory gene expression through the phosphatidylinositol 3-kinase pathway in vascular smooth muscle cells. J Biol Chem 277:36631–36639 [CrossRef]
    [Google Scholar]
  55. Shirakawa K., Maeda S., Gotoh T., Hayashi M., Shinomiya K., Ehata S., Nishimura R., Mori M., Onozaki K. other authors 2006; CCAAT/enhancer-binding protein homologous protein (CHOP) regulates osteoblast differentiation. Mol Cell Biol 26:6105–6116 [CrossRef]
    [Google Scholar]
  56. Simone C., Stiegler P., Bagella L., Pucci B., Bellan C., De Falco G., De Luca A., Guanti G., Puri P. L., Giordano A. 2002; Activation of MyoD-dependent transcription by cdk9/cyclin T2. Oncogene 21:4137–4148 [CrossRef]
    [Google Scholar]
  57. Spooner C. J., Guo X., Johnson P. F., Schwartz R. C. 2007; Differential roles of C/EBP beta regulatory domains in specifying MCP-1 and IL-6 transcription. Mol Immunol 44:1384–1392 [CrossRef]
    [Google Scholar]
  58. Wei P., Garber M. E., Fang S. M., Fischer W. H., Jones K. A. 1998; A novel CDK9-associated C-type cyclin interacts directly with HIV-1 Tat and mediates its high-affinity, loop-specific binding to TAR RNA. Cell 92:451–462 [CrossRef]
    [Google Scholar]
  59. Westendorf J. J., Yamamoto C. M., Lenny N., Downing J. R., Selsted M. E., Hiebert S. W. 1998; The t(8; 21) fusion product, AML-1-ETO, associates with C/EBP-alpha, inhibits C/EBP-alpha-dependent transcription, and blocks granulocytic differentiation. Mol Cell Biol 18:322–333
    [Google Scholar]
  60. Yik J. H., Chen R., Pezda A. C., Samford C. S., Zhou Q. 2004; A human immunodeficiency virus type 1 Tat-like arginine-rich RNA-binding domain is essential for HEXIM1 to inhibit RNA polymerase II transcription through 7SK snRNA-mediated inactivation of P-TEFb. Mol Cell Biol 24:5094–5105 [CrossRef]
    [Google Scholar]
  61. Yin M., Yang S. Q., Lin H. Z., Lane M. D., Chatterjee S., Diehl A. M. 1996; Tumor necrosis factor alpha promotes nuclear localization of cytokine-inducible CCAAT/enhancer binding protein isoforms in hepatocytes. J Biol Chem 271:17974–17978 [CrossRef]
    [Google Scholar]
  62. Yu W., Wang Y., Shaw C. A., Qin X. F., Rice A. P. 2006; Induction of the HIV-1 Tat co-factor cyclin T1 during monocyte differentiation is required for the regulated expression of a large portion of cellular mRNAs. Retrovirology 3:32 [CrossRef]
    [Google Scholar]
  63. Zhang Y., Nakata K., Weiden M., Rom W. N. 1995; Mycobacterium tuberculosis enhances human immunodeficiency virus-1 replication by transcriptional activation at the long terminal repeat. J Clin Invest 95:2324–2331 [CrossRef]
    [Google Scholar]
  64. Zhang W., Hirshberg M., McLaughlin S. H., Lazar G. A., Grossmann J. G., Nielsen P. R., Sobott F., Robinson C. V., Jackson S. E., Laue E. D. 2004; Biochemical and structural studies of the interaction of Cdc37 with Hsp90. J Mol Biol 340:891–907 [CrossRef]
    [Google Scholar]
  65. Zhu Y., Pe'ery T., Peng J., Ramanathan Y., Marshall N., Marshall T., Amendt B., Mathews M. B., Price D. H. 1997; Transcription elongation factor P-TEFb is required for HIV-1 tat transactivation in vitro. Genes Dev 11:2622–2632 [CrossRef]
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/jgv/10.1099/vir.0.82487-0
Loading
/content/journal/jgv/10.1099/vir.0.82487-0
Loading

Data & Media loading...

Most cited this month Most Cited RSS feed

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error