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. 1994 Jan 15;13(2):391–399. doi: 10.1002/j.1460-2075.1994.tb06273.x

Transcription initiation by RNA polymerase II does not require hydrolysis of the beta-gamma phosphoanhydride bond of ATP.

H T Timmers 1
PMCID: PMC394820  PMID: 8313884

Abstract

When transcription by RNA polymerase II from the major-late (ML) promoter was studied with purified basal transcription factors, it was observed that transcription from negatively-supercoiled ML templates did not require transcription factor IIH (TFIIH). Addition of the basal factor TFIIE was highly stimulatory, but not absolutely required for this reaction. In contrast, transcription from relaxed or linear ML templates required both TFIIE and TFIIH. Adenylylimidodiphosphate (AMP-PNP), an ATP analog with a non-hydrolyzable beta-gamma phosphoanhydride bond, could support RNA synthesis from supercoiled templates, but not from linear templates. Since AMP-PNP cannot act as a cofactor for the DNA helicase activity of TFIIH, this finding independently supported the conclusion that TFIIH is not required for transcription of negatively-supercoiled templates. Taken together, these data indicate that the ATP-dependent step in transcription initiation by RNA polymerase II is caused by a requirement for the ATP-dependent helicase activity of the basal factor TFIIH. The experiments also show that transcription initiation by RNA polymerase II does not require hydrolysis of the beta-gamma phosphoanhydride bond of ATP per se.

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Selected References

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

  1. Bunick D., Zandomeni R., Ackerman S., Weinmann R. Mechanism of RNA polymerase II--specific initiation of transcription in vitro: ATP requirement and uncapped runoff transcripts. Cell. 1982 Jul;29(3):877–886. doi: 10.1016/0092-8674(82)90449-4. [DOI] [PubMed] [Google Scholar]
  2. Buratowski S. DNA repair and transcription: the helicase connection. Science. 1993 Apr 2;260(5104):37–38. doi: 10.1126/science.8465198. [DOI] [PubMed] [Google Scholar]
  3. Buratowski S., Sopta M., Greenblatt J., Sharp P. A. RNA polymerase II-associated proteins are required for a DNA conformation change in the transcription initiation complex. Proc Natl Acad Sci U S A. 1991 Sep 1;88(17):7509–7513. doi: 10.1073/pnas.88.17.7509. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Carthew R. W., Samuels M., Sharp P. A. Formation of transcription preinitiation complexes with an amanitin-resistant RNA polymerase II. J Biol Chem. 1988 Nov 15;263(32):17128–17135. [PubMed] [Google Scholar]
  5. Challberg M. D., Kelly T. J., Jr Adenovirus DNA replication in vitro. Proc Natl Acad Sci U S A. 1979 Feb;76(2):655–659. doi: 10.1073/pnas.76.2.655. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Chesnut J. D., Stephens J. H., Dahmus M. E. The interaction of RNA polymerase II with the adenovirus-2 major late promoter is precluded by phosphorylation of the C-terminal domain of subunit IIa. J Biol Chem. 1992 May 25;267(15):10500–10506. [PubMed] [Google Scholar]
  7. Conaway J. W., Conaway R. C. A multisubunit transcription factor essential for accurate initiation by RNA polymerase II. J Biol Chem. 1989 Feb 5;264(4):2357–2362. [PubMed] [Google Scholar]
  8. Conaway J. W., Conaway R. C. An RNA polymerase II transcription factor shares functional properties with Escherichia coli sigma 70. Science. 1990 Jun 22;248(4962):1550–1553. doi: 10.1126/science.2193400. [DOI] [PubMed] [Google Scholar]
  9. Conaway J. W., Conaway R. C. Initiation of eukaryotic messenger RNA synthesis. J Biol Chem. 1991 Sep 25;266(27):17721–17724. [PubMed] [Google Scholar]
  10. Conaway R. C., Conaway J. W. ATP activates transcription initiation from promoters by RNA polymerase II in a reversible step prior to RNA synthesis. J Biol Chem. 1988 Feb 25;263(6):2962–2968. [PubMed] [Google Scholar]
  11. Conaway R. C., Conaway J. W. An RNA polymerase II transcription factor has an associated DNA-dependent ATPase (dATPase) activity strongly stimulated by the TATA region of promoters. Proc Natl Acad Sci U S A. 1989 Oct;86(19):7356–7360. doi: 10.1073/pnas.86.19.7356. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Fischer L., Gerard M., Chalut C., Lutz Y., Humbert S., Kanno M., Chambon P., Egly J. M. Cloning of the 62-kilodalton component of basic transcription factor BTF2. Science. 1992 Sep 4;257(5075):1392–1395. doi: 10.1126/science.1529339. [DOI] [PubMed] [Google Scholar]
  13. Flores O., Lu H., Reinberg D. Factors involved in specific transcription by mammalian RNA polymerase II. Identification and characterization of factor IIH. J Biol Chem. 1992 Feb 5;267(4):2786–2793. [PubMed] [Google Scholar]
  14. Gerard M., Fischer L., Moncollin V., Chipoulet J. M., Chambon P., Egly J. M. Purification and interaction properties of the human RNA polymerase B(II) general transcription factor BTF2. J Biol Chem. 1991 Nov 5;266(31):20940–20945. [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. Killeen M. T., Greenblatt J. F. The general transcription factor RAP30 binds to RNA polymerase II and prevents it from binding nonspecifically to DNA. Mol Cell Biol. 1992 Jan;12(1):30–37. doi: 10.1128/mcb.12.1.30. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Laybourn P. J., Dahmus M. E. Phosphorylation of RNA polymerase IIA occurs subsequent to interaction with the promoter and before the initiation of transcription. J Biol Chem. 1990 Aug 5;265(22):13165–13173. [PubMed] [Google Scholar]
  18. Leonard M. W., Patient R. K. Evidence for torsional stress in transcriptionally activated chromatin. Mol Cell Biol. 1991 Dec;11(12):6128–6138. doi: 10.1128/mcb.11.12.6128. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Lofquist A. K., Li H., Imboden M. A., Paule M. R. Promoter opening (melting) and transcription initiation by RNA polymerase I requires neither nucleotide beta,gamma hydrolysis nor protein phosphorylation. Nucleic Acids Res. 1993 Jul 11;21(14):3233–3238. doi: 10.1093/nar/21.14.3233. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Lu H., Flores O., Weinmann R., Reinberg D. The nonphosphorylated form of RNA polymerase II preferentially associates with the preinitiation complex. Proc Natl Acad Sci U S A. 1991 Nov 15;88(22):10004–10008. doi: 10.1073/pnas.88.22.10004. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Lu H., Zawel L., Fisher L., Egly J. M., Reinberg D. Human general transcription factor IIH phosphorylates the C-terminal domain of RNA polymerase II. Nature. 1992 Aug 20;358(6388):641–645. doi: 10.1038/358641a0. [DOI] [PubMed] [Google Scholar]
  22. Mitchell P. J., Tjian R. Transcriptional regulation in mammalian cells by sequence-specific DNA binding proteins. Science. 1989 Jul 28;245(4916):371–378. doi: 10.1126/science.2667136. [DOI] [PubMed] [Google Scholar]
  23. Mizutani M., Ohta T., Watanabe H., Handa H., Hirose S. Negative supercoiling of DNA facilitates an interaction between transcription factor IID and the fibroin gene promoter. Proc Natl Acad Sci U S A. 1991 Feb 1;88(3):718–722. doi: 10.1073/pnas.88.3.718. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Parvin J. D., Sharp P. A. DNA topology and a minimal set of basal factors for transcription by RNA polymerase II. Cell. 1993 May 7;73(3):533–540. doi: 10.1016/0092-8674(93)90140-l. [DOI] [PubMed] [Google Scholar]
  25. Parvin J. D., Timmers H. T., Sharp P. A. Promoter specificity of basal transcription factors. Cell. 1992 Mar 20;68(6):1135–1144. doi: 10.1016/0092-8674(92)90084-p. [DOI] [PubMed] [Google Scholar]
  26. Peterson M. G., Inostroza J., Maxon M. E., Flores O., Admon A., Reinberg D., Tjian R. Structure and functional properties of human general transcription factor IIE. Nature. 1991 Dec 5;354(6352):369–373. doi: 10.1038/354369a0. [DOI] [PubMed] [Google Scholar]
  27. Pognonec P., Kato H., Sumimoto H., Kretzschmar M., Roeder R. G. A quick procedure for purification of functional recombinant proteins over-expressed in E.coli. Nucleic Acids Res. 1991 Dec 11;19(23):6650–6650. doi: 10.1093/nar/19.23.6650. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Rappaport J., Weinmann R. Purine triphosphate beta-gamma bond hydrolysis requirements for RNA polymerase II transcription initiation and elongation. J Biol Chem. 1987 Dec 25;262(36):17510–17515. [PubMed] [Google Scholar]
  29. 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]
  30. Ryffel G. U., Kugler W., Wagner U., Kaling M. Liver cell specific gene transcription in vitro: the promoter elements HP1 and TATA box are necessary and sufficient to generate a liver-specific promoter. Nucleic Acids Res. 1989 Feb 11;17(3):939–953. doi: 10.1093/nar/17.3.939. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Samuels M., Fire A., Sharp P. A. Separation and characterization of factors mediating accurate transcription by RNA polymerase II. J Biol Chem. 1982 Dec 10;257(23):14419–14427. [PubMed] [Google Scholar]
  32. Sawadogo M., Roeder R. G. Energy requirement for specific transcription initiation by the human RNA polymerase II system. J Biol Chem. 1984 Apr 25;259(8):5321–5326. [PubMed] [Google Scholar]
  33. Sawadogo M., Roeder R. G. Factors involved in specific transcription by human RNA polymerase II: analysis by a rapid and quantitative in vitro assay. Proc Natl Acad Sci U S A. 1985 Jul;82(13):4394–4398. doi: 10.1073/pnas.82.13.4394. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Sawadogo M., Sentenac A. RNA polymerase B (II) and general transcription factors. Annu Rev Biochem. 1990;59:711–754. doi: 10.1146/annurev.bi.59.070190.003431. [DOI] [PubMed] [Google Scholar]
  35. Schaeffer L., Roy R., Humbert S., Moncollin V., Vermeulen W., Hoeijmakers J. H., Chambon P., Egly J. M. DNA repair helicase: a component of BTF2 (TFIIH) basic transcription factor. Science. 1993 Apr 2;260(5104):58–63. doi: 10.1126/science.8465201. [DOI] [PubMed] [Google Scholar]
  36. Schultz M. C., Brill S. J., Ju Q., Sternglanz R., Reeder R. H. Topoisomerases and yeast rRNA transcription: negative supercoiling stimulates initiation and topoisomerase activity is required for elongation. Genes Dev. 1992 Jul;6(7):1332–1341. doi: 10.1101/gad.6.7.1332. [DOI] [PubMed] [Google Scholar]
  37. Serizawa H., Conaway J. W., Conaway R. C. Phosphorylation of C-terminal domain of RNA polymerase II is not required in basal transcription. Nature. 1993 May 27;363(6427):371–374. doi: 10.1038/363371a0. [DOI] [PubMed] [Google Scholar]
  38. Serizawa H., Conaway R. C., Conaway J. W. A carboxyl-terminal-domain kinase associated with RNA polymerase II transcription factor delta from rat liver. Proc Natl Acad Sci U S A. 1992 Aug 15;89(16):7476–7480. doi: 10.1073/pnas.89.16.7476. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Sinden R. R., Carlson J. O., Pettijohn D. E. Torsional tension in the DNA double helix measured with trimethylpsoralen in living E. coli cells: analogous measurements in insect and human cells. Cell. 1980 Oct;21(3):773–783. doi: 10.1016/0092-8674(80)90440-7. [DOI] [PubMed] [Google Scholar]
  40. Szentirmay M. N., Sawadogo M. Transcription factor requirement for multiple rounds of initiation by human RNA polymerase II. Proc Natl Acad Sci U S A. 1991 Dec 1;88(23):10691–10695. doi: 10.1073/pnas.88.23.10691. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Timmers H. T., Sharp P. A. The mammalian TFIID protein is present in two functionally distinct complexes. Genes Dev. 1991 Nov;5(11):1946–1956. doi: 10.1101/gad.5.11.1946. [DOI] [PubMed] [Google Scholar]
  42. 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]
  43. Wang W., Carey M., Gralla J. D. Polymerase II promoter activation: closed complex formation and ATP-driven start site opening. Science. 1992 Jan 24;255(5043):450–453. doi: 10.1126/science.1310361. [DOI] [PubMed] [Google Scholar]
  44. Zawel L., Reinberg D. Advances in RNA polymerase II transcription. Curr Opin Cell Biol. 1992 Jun;4(3):488–495. doi: 10.1016/0955-0674(92)90016-6. [DOI] [PubMed] [Google Scholar]

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