Skip to main content
NCBI home page
The EMBO Journal logoLink to The EMBO Journal
. 1996 Dec 16;15(24):7079–7087.

The transcriptional corepressor DSP1 inhibits activated transcription by disrupting TFIIA-TBP complex formation.

N C Kirov 1, P M Lieberman 1, C Rushlow 1
PMCID: PMC452533  PMID: 9003783

Abstract

Transcriptional repression of eukaryotic genes is essential for many cellular and developmental processes, yet the precise mechanisms of repression remain poorly understood. The Dorsal Switch Protein (DSP1) was identified in a genetic screen for activities which convert Dorsal into a transcriptional repressor. DSP1 shares structural homology with the HMG-1/2 family and inhibits activation by the rel transcription factors Dorsal and NF-kappaB in transfection studies. Here we investigate the mechanism of transcriptional repression by DSP1. We found that DSP1 protein can act as a potent transcriptional repressor for multiple activator families in vitro and in transfection studies. DSP1 bound directly to the TATA binding protein (TBP), and formed a stable ternary complex with TBP bound to DNA. DSP1 preferentially disrupted the DNA binding of TBP complexes containing TFIIA and displaced TFIIA from binding to TBP. Consistent with the inhibition of TFIIA-bound complexes, DSP1 was shown to inhibit activated but not basal transcription reactions in vitro. The ability of DSP1 to interact with TBP and to repress transcription was mapped to the carboxy-terminal domain which contains two HMG boxes. Our results support the model that DSP1 represses activated transcription by interfering with the binding of TFIIA, a general transcription factor implicated in activated transcription pathways.

Full text

PDF
7079

Images in this article

7081Image
on p.7081
7082Image
on p.7082
7082Image
on p.7082
7083Image
on p.7083
7083Image
on p.7083
7084Image
on p.7084
7084Image
on p.7084
7084Image
on p.7084

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Auble D. T., Hansen K. E., Mueller C. G., Lane W. S., Thorner J., Hahn S. Mot1, a global repressor of RNA polymerase II transcription, inhibits TBP binding to DNA by an ATP-dependent mechanism. Genes Dev. 1994 Aug 15;8(16):1920–1934. doi: 10.1101/gad.8.16.1920. [DOI] [PubMed] [Google Scholar]
  2. Ayer D. E., Lawrence Q. A., Eisenman R. N. Mad-Max transcriptional repression is mediated by ternary complex formation with mammalian homologs of yeast repressor Sin3. Cell. 1995 Mar 10;80(5):767–776. doi: 10.1016/0092-8674(95)90355-0. [DOI] [PubMed] [Google Scholar]
  3. Bazett-Jones D. P., Leblanc B., Herfort M., Moss T. Short-range DNA looping by the Xenopus HMG-box transcription factor, xUBF. Science. 1994 May 20;264(5162):1134–1137. doi: 10.1126/science.8178172. [DOI] [PubMed] [Google Scholar]
  4. Buratowski S., Hahn S., Guarente L., Sharp P. A. Five intermediate complexes in transcription initiation by RNA polymerase II. Cell. 1989 Feb 24;56(4):549–561. doi: 10.1016/0092-8674(89)90578-3. [DOI] [PubMed] [Google Scholar]
  5. Buratowski S. The basics of basal transcription by RNA polymerase II. Cell. 1994 Apr 8;77(1):1–3. doi: 10.1016/0092-8674(94)90226-7. [DOI] [PubMed] [Google Scholar]
  6. Carey M. F. Transcriptional activation. A holistic view of the complex. Curr Biol. 1995 Sep 1;5(9):1003–1005. doi: 10.1016/s0960-9822(95)00201-6. [DOI] [PubMed] [Google Scholar]
  7. Carey M., Kolman J., Katz D. A., Gradoville L., Barberis L., Miller G. Transcriptional synergy by the Epstein-Barr virus transactivator ZEBRA. J Virol. 1992 Aug;66(8):4803–4813. doi: 10.1128/jvi.66.8.4803-4813.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Chasman D. I., Leatherwood J., Carey M., Ptashne M., Kornberg R. D. Activation of yeast polymerase II transcription by herpesvirus VP16 and GAL4 derivatives in vitro. Mol Cell Biol. 1989 Nov;9(11):4746–4749. doi: 10.1128/mcb.9.11.4746. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Conaway R. C., Conaway J. W. General initiation factors for RNA polymerase II. Annu Rev Biochem. 1993;62:161–190. doi: 10.1146/annurev.bi.62.070193.001113. [DOI] [PubMed] [Google Scholar]
  10. Cortes P., Flores O., Reinberg D. Factors involved in specific transcription by mammalian RNA polymerase II: purification and analysis of transcription factor IIA and identification of transcription factor IIJ. Mol Cell Biol. 1992 Jan;12(1):413–421. doi: 10.1128/mcb.12.1.413. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Fondell J. D., Roy A. L., Roeder R. G. Unliganded thyroid hormone receptor inhibits formation of a functional preinitiation complex: implications for active repression. Genes Dev. 1993 Jul;7(7B):1400–1410. doi: 10.1101/gad.7.7b.1400. [DOI] [PubMed] [Google Scholar]
  12. Ge H., Roeder R. G. The high mobility group protein HMG1 can reversibly inhibit class II gene transcription by interaction with the TATA-binding protein. J Biol Chem. 1994 Jun 24;269(25):17136–17140. [PubMed] [Google Scholar]
  13. Goodrich J. A., Tjian R. TBP-TAF complexes: selectivity factors for eukaryotic transcription. Curr Opin Cell Biol. 1994 Jun;6(3):403–409. doi: 10.1016/0955-0674(94)90033-7. [DOI] [PubMed] [Google Scholar]
  14. Grosschedl R., Giese K., Pagel J. HMG domain proteins: architectural elements in the assembly of nucleoprotein structures. Trends Genet. 1994 Mar;10(3):94–100. doi: 10.1016/0168-9525(94)90232-1. [DOI] [PubMed] [Google Scholar]
  15. Ha I., Lane W. S., Reinberg D. Cloning of a human gene encoding the general transcription initiation factor IIB. Nature. 1991 Aug 22;352(6337):689–695. doi: 10.1038/352689a0. [DOI] [PubMed] [Google Scholar]
  16. Han K., Manley J. L. Functional domains of the Drosophila Engrailed protein. EMBO J. 1993 Jul;12(7):2723–2733. doi: 10.1002/j.1460-2075.1993.tb05934.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Han K., Manley J. L. Transcriptional repression by the Drosophila even-skipped protein: definition of a minimal repression domain. Genes Dev. 1993 Mar;7(3):491–503. doi: 10.1101/gad.7.3.491. [DOI] [PubMed] [Google Scholar]
  18. Hanna-Rose W., Hansen U. Active repression mechanisms of eukaryotic transcription repressors. Trends Genet. 1996 Jun;12(6):229–234. doi: 10.1016/0168-9525(96)10022-6. [DOI] [PubMed] [Google Scholar]
  19. Higuchi R., Krummel B., Saiki R. K. A general method of in vitro preparation and specific mutagenesis of DNA fragments: study of protein and DNA interactions. Nucleic Acids Res. 1988 Aug 11;16(15):7351–7367. doi: 10.1093/nar/16.15.7351. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Imbalzano A. N., Zaret K. S., Kingston R. E. Transcription factor (TF) IIB and TFIIA can independently increase the affinity of the TATA-binding protein for DNA. J Biol Chem. 1994 Mar 18;269(11):8280–8286. [PubMed] [Google Scholar]
  21. Inostroza J. A., Mermelstein F. H., Ha I., Lane W. S., Reinberg D. Dr1, a TATA-binding protein-associated phosphoprotein and inhibitor of class II gene transcription. Cell. 1992 Aug 7;70(3):477–489. doi: 10.1016/0092-8674(92)90172-9. [DOI] [PubMed] [Google Scholar]
  22. Jiang J., Cai H., Zhou Q., Levine M. Conversion of a dorsal-dependent silencer into an enhancer: evidence for dorsal corepressors. EMBO J. 1993 Aug;12(8):3201–3209. doi: 10.1002/j.1460-2075.1993.tb05989.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Johnson A. D. The price of repression. Cell. 1995 Jun 2;81(5):655–658. doi: 10.1016/0092-8674(95)90524-3. [DOI] [PubMed] [Google Scholar]
  24. Kao C. C., Lieberman P. M., Schmidt M. C., Zhou Q., Pei R., Berk A. J. Cloning of a transcriptionally active human TATA binding factor. Science. 1990 Jun 29;248(4963):1646–1650. doi: 10.1126/science.2194289. [DOI] [PubMed] [Google Scholar]
  25. Kerppola T. K., Luk D., Curran T. Fos is a preferential target of glucocorticoid receptor inhibition of AP-1 activity in vitro. Mol Cell Biol. 1993 Jun;13(6):3782–3791. doi: 10.1128/mcb.13.6.3782. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Kim T. K., Roeder R. G. Transcriptional activation in yeast by the proline-rich activation domain of human CTF1. J Biol Chem. 1993 Oct 5;268(28):20866–20869. [PubMed] [Google Scholar]
  27. Kirov N., Zhelnin L., Shah J., Rushlow C. Conversion of a silencer into an enhancer: evidence for a co-repressor in dorsal-mediated repression in Drosophila. EMBO J. 1993 Aug;12(8):3193–3199. doi: 10.1002/j.1460-2075.1993.tb05988.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Koleske A. J., Young R. A. The RNA polymerase II holoenzyme and its implications for gene regulation. Trends Biochem Sci. 1995 Mar;20(3):113–116. doi: 10.1016/s0968-0004(00)88977-x. [DOI] [PubMed] [Google Scholar]
  29. Kuchin S., Yeghiayan P., Carlson M. Cyclin-dependent protein kinase and cyclin homologs SSN3 and SSN8 contribute to transcriptional control in yeast. Proc Natl Acad Sci U S A. 1995 Apr 25;92(9):4006–4010. doi: 10.1073/pnas.92.9.4006. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Lai J. S., Herr W. Ethidium bromide provides a simple tool for identifying genuine DNA-independent protein associations. Proc Natl Acad Sci U S A. 1992 Aug 1;89(15):6958–6962. doi: 10.1073/pnas.89.15.6958. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Landsman D., Bustin M. A signature for the HMG-1 box DNA-binding proteins. Bioessays. 1993 Aug;15(8):539–546. doi: 10.1002/bies.950150807. [DOI] [PubMed] [Google Scholar]
  32. Lehming N., Thanos D., Brickman J. M., Ma J., Maniatis T., Ptashne M. An HMG-like protein that can switch a transcriptional activator to a repressor. Nature. 1994 Sep 8;371(6493):175–179. doi: 10.1038/371175a0. [DOI] [PubMed] [Google Scholar]
  33. Lieberman P. M., Berk A. J. A mechanism for TAFs in transcriptional activation: activation domain enhancement of TFIID-TFIIA--promoter DNA complex formation. Genes Dev. 1994 May 1;8(9):995–1006. doi: 10.1101/gad.8.9.995. [DOI] [PubMed] [Google Scholar]
  34. Lieberman P. M., Berk A. J. The Zta trans-activator protein stabilizes TFIID association with promoter DNA by direct protein-protein interaction. Genes Dev. 1991 Dec;5(12B):2441–2454. doi: 10.1101/gad.5.12b.2441. [DOI] [PubMed] [Google Scholar]
  35. Lin Y. S., Carey M. F., Ptashne M., Green M. R. GAL4 derivatives function alone and synergistically with mammalian activators in vitro. Cell. 1988 Aug 26;54(5):659–664. doi: 10.1016/s0092-8674(88)80010-2. [DOI] [PubMed] [Google Scholar]
  36. Lin Y. S., Carey M. F., Ptashne M., Green M. R. GAL4 derivatives function alone and synergistically with mammalian activators in vitro. Cell. 1988 Aug 26;54(5):659–664. doi: 10.1016/s0092-8674(88)80010-2. [DOI] [PubMed] [Google Scholar]
  37. Lin Y. S., Ha I., Maldonado E., Reinberg D., Green M. R. Binding of general transcription factor TFIIB to an acidic activating region. Nature. 1991 Oct 10;353(6344):569–571. doi: 10.1038/353569a0. [DOI] [PubMed] [Google Scholar]
  38. Maldonado E., Ha I., Cortes P., Weis L., Reinberg D. Factors involved in specific transcription by mammalian RNA polymerase II: role of transcription factors IIA, IID, and IIB during formation of a transcription-competent complex. Mol Cell Biol. 1990 Dec;10(12):6335–6347. doi: 10.1128/mcb.10.12.6335. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Meisterernst M., Roy A. L., Lieu H. M., Roeder R. G. Activation of class II gene transcription by regulatory factors is potentiated by a novel activity. Cell. 1991 Sep 6;66(5):981–993. doi: 10.1016/0092-8674(91)90443-3. [DOI] [PubMed] [Google Scholar]
  40. Merino A., Madden K. R., Lane W. S., Champoux J. J., Reinberg D. DNA topoisomerase I is involved in both repression and activation of transcription. Nature. 1993 Sep 16;365(6443):227–232. doi: 10.1038/365227a0. [DOI] [PubMed] [Google Scholar]
  41. Ozer J., Moore P. A., Bolden A. H., Lee A., Rosen C. A., Lieberman P. M. Molecular cloning of the small (gamma) subunit of human TFIIA reveals functions critical for activated transcription. Genes Dev. 1994 Oct 1;8(19):2324–2335. doi: 10.1101/gad.8.19.2324. [DOI] [PubMed] [Google Scholar]
  42. Ptashne M., Gann A. A. Activators and targets. Nature. 1990 Jul 26;346(6282):329–331. doi: 10.1038/346329a0. [DOI] [PubMed] [Google Scholar]
  43. Roeder R. G. The complexities of eukaryotic transcription initiation: regulation of preinitiation complex assembly. Trends Biochem Sci. 1991 Nov;16(11):402–408. doi: 10.1016/0968-0004(91)90164-q. [DOI] [PubMed] [Google Scholar]
  44. Sauer F., Fondell J. D., Ohkuma Y., Roeder R. G., Jäckle H. Control of transcription by Krüppel through interactions with TFIIB and TFIIE beta. Nature. 1995 May 11;375(6527):162–164. doi: 10.1038/375162a0. [DOI] [PubMed] [Google Scholar]
  45. Schreiber-Agus N., Chin L., Chen K., Torres R., Rao G., Guida P., Skoultchi A. I., DePinho R. A. An amino-terminal domain of Mxi1 mediates anti-Myc oncogenic activity and interacts with a homolog of the yeast transcriptional repressor SIN3. Cell. 1995 Mar 10;80(5):777–786. doi: 10.1016/0092-8674(95)90356-9. [DOI] [PubMed] [Google Scholar]
  46. Seto E., Usheva A., Zambetti G. P., Momand J., Horikoshi N., Weinmann R., Levine A. J., Shenk T. Wild-type p53 binds to the TATA-binding protein and represses transcription. Proc Natl Acad Sci U S A. 1992 Dec 15;89(24):12028–12032. doi: 10.1073/pnas.89.24.12028. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Shykind B. M., Kim J., Sharp P. A. Activation of the TFIID-TFIIA complex with HMG-2. Genes Dev. 1995 Jun 1;9(11):1354–1365. doi: 10.1101/gad.9.11.1354. [DOI] [PubMed] [Google Scholar]
  48. Smith D. B., Johnson K. S. Single-step purification of polypeptides expressed in Escherichia coli as fusions with glutathione S-transferase. Gene. 1988 Jul 15;67(1):31–40. doi: 10.1016/0378-1119(88)90005-4. [DOI] [PubMed] [Google Scholar]
  49. Song W., Treich I., Qian N., Kuchin S., Carlson M. SSN genes that affect transcriptional repression in Saccharomyces cerevisiae encode SIN4, ROX3, and SRB proteins associated with RNA polymerase II. Mol Cell Biol. 1996 Jan;16(1):115–120. doi: 10.1128/mcb.16.1.115. [DOI] [PMC free article] [PubMed] [Google Scholar]
  50. Stargell L. A., Struhl K. The TBP-TFIIA interaction in the response to acidic activators in vivo. Science. 1995 Jul 7;269(5220):75–78. doi: 10.1126/science.7604282. [DOI] [PubMed] [Google Scholar]
  51. Stelzer G., Goppelt A., Lottspeich F., Meisterernst M. Repression of basal transcription by HMG2 is counteracted by TFIIH-associated factors in an ATP-dependent process. Mol Cell Biol. 1994 Jul;14(7):4712–4721. doi: 10.1128/mcb.14.7.4712. [DOI] [PMC free article] [PubMed] [Google Scholar]
  52. Sun X., Ma D., Sheldon M., Yeung K., Reinberg D. Reconstitution of human TFIIA activity from recombinant polypeptides: a role in TFIID-mediated transcription. Genes Dev. 1994 Oct 1;8(19):2336–2348. doi: 10.1101/gad.8.19.2336. [DOI] [PubMed] [Google Scholar]
  53. Tjian R., Maniatis T. Transcriptional activation: a complex puzzle with few easy pieces. Cell. 1994 Apr 8;77(1):5–8. doi: 10.1016/0092-8674(94)90227-5. [DOI] [PubMed] [Google Scholar]
  54. Tyree C. M., George C. P., Lira-DeVito L. M., Wampler S. L., Dahmus M. E., Zawel L., Kadonaga J. T. Identification of a minimal set of proteins that is sufficient for accurate initiation of transcription by RNA polymerase II. Genes Dev. 1993 Jul;7(7A):1254–1265. doi: 10.1101/gad.7.7a.1254. [DOI] [PubMed] [Google Scholar]
  55. Wahi M., Johnson A. D. Identification of genes required for alpha 2 repression in Saccharomyces cerevisiae. Genetics. 1995 May;140(1):79–90. doi: 10.1093/genetics/140.1.79. [DOI] [PMC free article] [PubMed] [Google Scholar]
  56. Wang W., Gralla J. D., Carey M. The acidic activator GAL4-AH can stimulate polymerase II transcription by promoting assembly of a closed complex requiring TFIID and TFIIA. Genes Dev. 1992 Sep;6(9):1716–1727. doi: 10.1101/gad.6.9.1716. [DOI] [PubMed] [Google Scholar]
  57. Weintraub S. J., Chow K. N., Luo R. X., Zhang S. H., He S., Dean D. C. Mechanism of active transcriptional repression by the retinoblastoma protein. Nature. 1995 Jun 29;375(6534):812–815. doi: 10.1038/375812a0. [DOI] [PubMed] [Google Scholar]
  58. Wolffe A. P. Architectural transcription factors. Science. 1994 May 20;264(5162):1100–1101. doi: 10.1126/science.8178167. [DOI] [PubMed] [Google Scholar]
  59. Yokomori K., Zeidler M. P., Chen J. L., Verrijzer C. P., Mlodzik M., Tjian R. Drosophila TFIIA directs cooperative DNA binding with TBP and mediates transcriptional activation. Genes Dev. 1994 Oct 1;8(19):2313–2323. doi: 10.1101/gad.8.19.2313. [DOI] [PubMed] [Google Scholar]
  60. Zawel L., Reinberg D. Initiation of transcription by RNA polymerase II: a multi-step process. Prog Nucleic Acid Res Mol Biol. 1993;44:67–108. doi: 10.1016/s0079-6603(08)60217-2. [DOI] [PubMed] [Google Scholar]
  61. Zhou Q., Lieberman P. M., Boyer T. G., Berk A. J. Holo-TFIID supports transcriptional stimulation by diverse activators and from a TATA-less promoter. Genes Dev. 1992 Oct;6(10):1964–1974. doi: 10.1101/gad.6.10.1964. [DOI] [PubMed] [Google Scholar]

Articles from The EMBO Journal are provided here courtesy of Nature Publishing Group

RESOURCES