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J Virol. Oct 1993; 67(10): 6224–6233.
PMCID: PMC238044

In vitro and in vivo binding of human immunodeficiency virus type 1 Tat protein and Sp1 transcription factor.

Abstract

Recent genetic experiments have suggested that tat transactivation of the human immunodeficiency virus type 1 (HIV-1) long terminal repeat requires functional upstream enhancer sequences--Sp1 sites, in particular. In these experiments, HeLa cell nuclear extracts were passed over affinity matrices containing chemically synthesized or bacterially expressed HIV-1 Tat. Assay of material that bound to and eluted from the Tat matrices revealed the presence of the Sp1 transcription factor. Other transcription factors (Oct and NF-kappa B) also bound to Tat matrices but with less efficiency--in parallel with the lower capacities of these binding motifs to confer Tat responsiveness on a basal HIV-1 promoter compared with Sp1 sites. Passage of nuclear extracts over matrices containing other neutral proteins, including bovine serum albumin, ovalbumin, and lysozyme, revealed no or reduced binding. Cross-linking experiments indicated that the purified Sp1 and Tat proteins can form multimeric complexes in the absence of other proteins. The region of Tat responsible for Sp1 binding was localized to a region encompassing residues 30 to 62. Immunoprecipitation experiments with HIV-1-infected T lymphocytes indicated coimmunoprecipitation of Tat and Sp1. These experiments extend previous genetic experiments and suggest a direct interaction between Tat and Sp1 during transactivation.

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These references are in PubMed. This may not be the complete list of references from this article.
  • Adachi A, Gendelman HE, Koenig S, Folks T, Willey R, Rabson A, Martin MA. Production of acquired immunodeficiency syndrome-associated retrovirus in human and nonhuman cells transfected with an infectious molecular clone. J Virol. 1986 Aug;59(2):284–291. [PMC free article] [PubMed]
  • Allet B, Payton M, Mattaliano RJ, Gronenborn AM, Clore GM, Wingfield PT. Purification and characterization of the DNA-binding protein Ner of bacteriophage Mu. Gene. 1988 May 30;65(2):259–268. [PubMed]
  • Arya SK, Guo C, Josephs SF, Wong-Staal F. Trans-activator gene of human T-lymphotropic virus type III (HTLV-III). Science. 1985 Jul 5;229(4708):69–73. [PubMed]
  • Atchison ML, Delmas V, Perry RP. A novel upstream element compensates for an ineffectual octamer motif in an immunoglobulin V kappa promoter. EMBO J. 1990 Oct;9(10):3109–3117. [PMC free article] [PubMed]
  • Berkhout B, Gatignol A, Silver J, Jeang KT. Efficient trans-activation by the HIV-2 Tat protein requires a duplicated TAR RNA structure. Nucleic Acids Res. 1990 Apr 11;18(7):1839–1846. [PMC free article] [PubMed]
  • Berkhout B, Jeang KT. Functional roles for the TATA promoter and enhancers in basal and Tat-induced expression of the human immunodeficiency virus type 1 long terminal repeat. J Virol. 1992 Jan;66(1):139–149. [PMC free article] [PubMed]
  • Berkhout B, Silverman RH, Jeang KT. Tat trans-activates the human immunodeficiency virus through a nascent RNA target. Cell. 1989 Oct 20;59(2):273–282. [PubMed]
  • Bohan CA, Kashanchi F, Ensoli B, Buonaguro L, Boris-Lawrie KA, Brady JN. Analysis of Tat transactivation of human immunodeficiency virus transcription in vitro. Gene Expr. 1992;2(4):391–407. [PubMed]
  • Brady J, Jeang KT, Duvall J, Khoury G. Identification of p40x-responsive regulatory sequences within the human T-cell leukemia virus type I long terminal repeat. J Virol. 1987 Jul;61(7):2175–2181. [PMC free article] [PubMed]
  • Briggs MR, Kadonaga JT, Bell SP, Tjian R. Purification and biochemical characterization of the promoter-specific transcription factor, Sp1. Science. 1986 Oct 3;234(4772):47–52. [PubMed]
  • Calnan BJ, Biancalana S, Hudson D, Frankel AD. Analysis of arginine-rich peptides from the HIV Tat protein reveals unusual features of RNA-protein recognition. Genes Dev. 1991 Feb;5(2):201–210. [PubMed]
  • Chun R, Glabe CG, Fan H. Chemical synthesis of biologically active tat trans-activating protein of human immunodeficiency virus type 1. J Virol. 1990 Jun;64(6):3074–3077. [PMC free article] [PubMed]
  • Cordingley MG, LaFemina RL, Callahan PL, Condra JH, Sardana VV, Graham DJ, Nguyen TM, LeGrow K, Gotlib L, Schlabach AJ, et al. Sequence-specific interaction of Tat protein and Tat peptides with the transactivation-responsive sequence element of human immunodeficiency virus type 1 in vitro. Proc Natl Acad Sci U S A. 1990 Nov;87(22):8985–8989. [PMC free article] [PubMed]
  • Courey AJ, Tjian R. Analysis of Sp1 in vivo reveals multiple transcriptional domains, including a novel glutamine-rich activation motif. Cell. 1988 Dec 2;55(5):887–898. [PubMed]
  • Desai K, Loewenstein PM, Green M. Isolation of a cellular protein that binds to the human immunodeficiency virus Tat protein and can potentiate transactivation of the viral promoter. Proc Natl Acad Sci U S A. 1991 Oct 15;88(20):8875–8879. [PMC free article] [PubMed]
  • Dignam JD, Lebovitz RM, Roeder RG. Accurate transcription initiation by RNA polymerase II in a soluble extract from isolated mammalian nuclei. Nucleic Acids Res. 1983 Mar 11;11(5):1475–1489. [PMC free article] [PubMed]
  • Dingwall C, Ernberg I, Gait MJ, Green SM, Heaphy S, Karn J, Lowe AD, Singh M, Skinner MA, Valerio R. Human immunodeficiency virus 1 tat protein binds trans-activation-responsive region (TAR) RNA in vitro. Proc Natl Acad Sci U S A. 1989 Sep;86(18):6925–6929. [PMC free article] [PubMed]
  • Dynan WS, Tjian R. Isolation of transcription factors that discriminate between different promoters recognized by RNA polymerase II. Cell. 1983 Mar;32(3):669–680. [PubMed]
  • Dynan WS, Tjian R. The promoter-specific transcription factor Sp1 binds to upstream sequences in the SV40 early promoter. Cell. 1983 Nov;35(1):79–87. [PubMed]
  • Dynlacht BD, Hoey T, Tjian R. Isolation of coactivators associated with the TATA-binding protein that mediate transcriptional activation. Cell. 1991 Aug 9;66(3):563–576. [PubMed]
  • Feinberg MB, Baltimore D, Frankel AD. The role of Tat in the human immunodeficiency virus life cycle indicates a primary effect on transcriptional elongation. Proc Natl Acad Sci U S A. 1991 May 1;88(9):4045–4049. [PMC free article] [PubMed]
  • Fisher AG, Feinberg MB, Josephs SF, Harper ME, Marselle LM, Reyes G, Gonda MA, Aldovini A, Debouk C, Gallo RC, et al. The trans-activator gene of HTLV-III is essential for virus replication. Nature. 320(6060):367–371. [PubMed]
  • Frankel AD, Bredt DS, Pabo CO. Tat protein from human immunodeficiency virus forms a metal-linked dimer. Science. 1988 Apr 1;240(4848):70–73. [PubMed]
  • Garcia JA, Harrich D, Soultanakis E, Wu F, Mitsuyasu R, Gaynor RB. Human immunodeficiency virus type 1 LTR TATA and TAR region sequences required for transcriptional regulation. EMBO J. 1989 Mar;8(3):765–778. [PMC free article] [PubMed]
  • Garcia JA, Wu FK, Mitsuyasu R, Gaynor RB. Interactions of cellular proteins involved in the transcriptional regulation of the human immunodeficiency virus. EMBO J. 1987 Dec 1;6(12):3761–3770. [PMC free article] [PubMed]
  • Gatignol A, Buckler-White A, Berkhout B, Jeang KT. Characterization of a human TAR RNA-binding protein that activates the HIV-1 LTR. Science. 1991 Mar 29;251(5001):1597–1600. [PubMed]
  • Gatignol A, Kumar A, Rabson A, Jeang KT. Identification of cellular proteins that bind to the human immunodeficiency virus type 1 trans-activation-responsive TAR element RNA. Proc Natl Acad Sci U S A. 1989 Oct;86(20):7828–7832. [PMC free article] [PubMed]
  • Gaynor R, Soultanakis E, Kuwabara M, Garcia J, Sigman DS. Specific binding of a HeLa cell nuclear protein to RNA sequences in the human immunodeficiency virus transactivating region. Proc Natl Acad Sci U S A. 1989 Jul;86(13):4858–4862. [PMC free article] [PubMed]
  • Gentz R, Chen CH, Rosen CA. Bioassay for trans-activation using purified human immunodeficiency virus tat-encoded protein: trans-activation requires mRNA synthesis. Proc Natl Acad Sci U S A. 1989 Feb;86(3):821–824. [PMC free article] [PubMed]
  • Greenblatt J. Roles of TFIID in transcriptional initiation by RNA polymerase II. Cell. 1991 Sep 20;66(6):1067–1070. [PubMed]
  • Harrich D, Garcia J, Wu F, Mitsuyasu R, Gonazalez J, Gaynor R. Role of SP1-binding domains in in vivo transcriptional regulation of the human immunodeficiency virus type 1 long terminal repeat. J Virol. 1989 Jun;63(6):2585–2591. [PMC free article] [PubMed]
  • Horikoshi N, Maguire K, Kralli A, Maldonado E, Reinberg D, Weinmann R. Direct interaction between adenovirus E1A protein and the TATA box binding transcription factor IID. Proc Natl Acad Sci U S A. 1991 Jun 15;88(12):5124–5128. [PMC free article] [PubMed]
  • Ingles CJ, Shales M, Cress WD, Triezenberg SJ, Greenblatt J. Reduced binding of TFIID to transcriptionally compromised mutants of VP16. Nature. 1991 Jun 13;351(6327):588–590. [PubMed]
  • Jeang KT, Boros I, Brady J, Radonovich M, Khoury G. Characterization of cellular factors that interact with the human T-cell leukemia virus type I p40x-responsive 21-base-pair sequence. J Virol. 1988 Dec;62(12):4499–4509. [PMC free article] [PubMed]
  • Jeang KT, Shank PR, Kumar A. Transcriptional activation of homologous viral long terminal repeats by the human immunodeficiency virus type 1 or the human T-cell leukemia virus type I tat proteins occurs in the absence of de novo protein synthesis. Proc Natl Acad Sci U S A. 1988 Nov;85(21):8291–8295. [PMC free article] [PubMed]
  • Jones KA, Kadonaga JT, Luciw PA, Tjian R. Activation of the AIDS retrovirus promoter by the cellular transcription factor, Sp1. Science. 1986 May 9;232(4751):755–759. [PubMed]
  • Jones KA, Luciw PA, Duchange N. Structural arrangements of transcription control domains within the 5'-untranslated leader regions of the HIV-1 and HIV-2 promoters. Genes Dev. 1988 Sep;2(9):1101–1114. [PubMed]
  • Kadonaga JT, Carner KR, Masiarz FR, Tjian R. Isolation of cDNA encoding transcription factor Sp1 and functional analysis of the DNA binding domain. Cell. 1987 Dec 24;51(6):1079–1090. [PubMed]
  • Kamine J, Subramanian T, Chinnadurai G. Sp1-dependent activation of a synthetic promoter by human immunodeficiency virus type 1 Tat protein. Proc Natl Acad Sci U S A. 1991 Oct 1;88(19):8510–8514. [PMC free article] [PubMed]
  • Kao SY, Calman AF, Luciw PA, Peterlin BM. Anti-termination of transcription within the long terminal repeat of HIV-1 by tat gene product. Nature. 1987 Dec 3;330(6147):489–493. [PubMed]
  • Laspia MF, Rice AP, Mathews MB. HIV-1 Tat protein increases transcriptional initiation and stabilizes elongation. Cell. 1989 Oct 20;59(2):283–292. [PubMed]
  • Leonard J, Parrott C, Buckler-White AJ, Turner W, Ross EK, Martin MA, Rabson AB. The NF-kappa B binding sites in the human immunodeficiency virus type 1 long terminal repeat are not required for virus infectivity. J Virol. 1989 Nov;63(11):4919–4924. [PMC free article] [PubMed]
  • Letovsky J, Dynan WS. Measurement of the binding of transcription factor Sp1 to a single GC box recognition sequence. Nucleic Acids Res. 1989 Apr 11;17(7):2639–2653. [PMC free article] [PubMed]
  • Lewin B. Commitment and activation at pol II promoters: a tail of protein-protein interactions. Cell. 1990 Jun 29;61(7):1161–1164. [PubMed]
  • Li R, Knight JD, Jackson SP, Tjian R, Botchan MR. Direct interaction between Sp1 and the BPV enhancer E2 protein mediates synergistic activation of transcription. Cell. 1991 May 3;65(3):493–505. [PubMed]
  • Lillie JW, Green MR. Gene transcription: activator's target in sight. Nature. 1989 Sep 28;341(6240):279–280. [PubMed]
  • Lin YS, Green MR. Mechanism of action of an acidic transcriptional activator in vitro. Cell. 1991 Mar 8;64(5):971–981. [PubMed]
  • Lin YS, Ha I, Maldonado E, Reinberg D, Green MR. Binding of general transcription factor TFIIB to an acidic activating region. Nature. 1991 Oct 10;353(6344):569–571. [PubMed]
  • Lu X, Welsh TM, Peterlin BM. The human immunodeficiency virus type 1 long terminal repeat specifies two different transcription complexes, only one of which is regulated by Tat. J Virol. 1993 Apr;67(4):1752–1760. [PMC free article] [PubMed]
  • Marciniak RA, Calnan BJ, Frankel AD, Sharp PA. HIV-1 Tat protein trans-activates transcription in vitro. Cell. 1990 Nov 16;63(4):791–802. [PubMed]
  • Marciniak RA, Sharp PA. HIV-1 Tat protein promotes formation of more-processive elongation complexes. EMBO J. 1991 Dec;10(13):4189–4196. [PMC free article] [PubMed]
  • Muesing MA, Smith DH, Capon DJ. Regulation of mRNA accumulation by a human immunodeficiency virus trans-activator protein. Cell. 1987 Feb 27;48(4):691–701. [PubMed]
  • Nabel G, Baltimore D. An inducible transcription factor activates expression of human immunodeficiency virus in T cells. Nature. 1987 Apr 16;326(6114):711–713. [PubMed]
  • Okamoto T, Wong-Staal F. Demonstration of virus-specific transcriptional activator(s) in cells infected with HTLV-III by an in vitro cell-free system. Cell. 1986 Oct 10;47(1):29–35. [PubMed]
  • Peterlin BM, Luciw PA, Barr PJ, Walker MD. Elevated levels of mRNA can account for the trans-activation of human immunodeficiency virus. Proc Natl Acad Sci U S A. 1986 Dec;83(24):9734–9738. [PMC free article] [PubMed]
  • Ptashne M, Gann AA. Activators and targets. Nature. 1990 Jul 26;346(6282):329–331. [PubMed]
  • Pugh BF, Tjian R. Mechanism of transcriptional activation by Sp1: evidence for coactivators. Cell. 1990 Jun 29;61(7):1187–1197. [PubMed]
  • Rice AP, Mathews MB. Transcriptional but not translational regulation of HIV-1 by the tat gene product. Nature. 1988 Apr 7;332(6164):551–553. [PubMed]
  • Rosen CA, Sodroski JG, Haseltine WA. The location of cis-acting regulatory sequences in the human T cell lymphotropic virus type III (HTLV-III/LAV) long terminal repeat. Cell. 1985 Jul;41(3):813–823. [PubMed]
  • Roy S, Parkin NT, Rosen C, Itovitch J, Sonenberg N. Structural requirements for trans activation of human immunodeficiency virus type 1 long terminal repeat-directed gene expression by tat: importance of base pairing, loop sequence, and bulges in the tat-responsive sequence. J Virol. 1990 Mar;64(3):1402–1406. [PMC free article] [PubMed]
  • Saffer JD, Jackson SP, Thurston SJ. SV40 stimulates expression of the transacting factor Sp1 at the mRNA level. Genes Dev. 1990 Apr;4(4):659–666. [PubMed]
  • Selby MJ, Peterlin BM. Trans-activation by HIV-1 Tat via a heterologous RNA binding protein. Cell. 1990 Aug 24;62(4):769–776. [PubMed]
  • Sharp PA. Gene transcription. TFIIB or not TFIIB? Nature. 1991 May 2;351(6321):16–18. [PubMed]
  • Sheline CT, Milocco LH, Jones KA. Two distinct nuclear transcription factors recognize loop and bulge residues of the HIV-1 TAR RNA hairpin. Genes Dev. 1991 Dec;5(12B):2508–2520. [PubMed]
  • Sodroski J, Patarca R, Rosen C, Wong-Staal F, Haseltine W. Location of the trans-activating region on the genome of human T-cell lymphotropic virus type III. Science. 1985 Jul 5;229(4708):74–77. [PubMed]
  • Southgate C, Zapp ML, Green MR. Activation of transcription by HIV-1 Tat protein tethered to nascent RNA through another protein. Nature. 1990 Jun 14;345(6276):640–642. [PubMed]
  • Southgate CD, Green MR. The HIV-1 Tat protein activates transcription from an upstream DNA-binding site: implications for Tat function. Genes Dev. 1991 Dec;5(12B):2496–2507. [PubMed]
  • Weeks KM, Ampe C, Schultz SC, Steitz TA, Crothers DM. Fragments of the HIV-1 Tat protein specifically bind TAR RNA. Science. 1990 Sep 14;249(4974):1281–1285. [PubMed]
  • Weeks KM, Crothers DM. RNA recognition by Tat-derived peptides: interaction in the major groove? Cell. 1991 Aug 9;66(3):577–588. [PubMed]
  • Wright CM, Felber BK, Paskalis H, Pavlakis GN. Expression and characterization of the trans-activator of HTLV-III/LAV virus. Science. 1986 Nov 21;234(4779):988–992. [PubMed]
  • Wu FK, Garcia JA, Harrich D, Gaynor RB. Purification of the human immunodeficiency virus type 1 enhancer and TAR binding proteins EBP-1 and UBP-1. EMBO J. 1988 Jul;7(7):2117–2130. [PMC free article] [PubMed]
  • Zimmermann K, Dobrovnik M, Ballaun C, Bevec D, Hauber J, Böhnlein E. trans-activation of the HIV-1 LTR by the HIV-1 Tat and HTLV-I Tax proteins is mediated by different cis-acting sequences. Virology. 1991 Jun;182(2):874–878. [PubMed]

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