What are the 4 steps in transcription initiation?

The progress of transcription bubbles during inhibition in vitro was followed in order to learn how RNA polymerase II begins transcription at the activated adenovirus E4 promoter. The issues addressed include the multiple roles of ATP, the potential effect of polymerase C-terminal domain phosphorylation, and the ability of polymerase to clear the promoter for reinitiation. The results lead to a three-step model for the transition from closed complex to elongation complex, two steps of which use ATP independently. In the first step, studied previously, ATP is hydrolyzed to open the DNA strands over the start site. In a second step, apparently independent of ATP, transcription bubbles move into the initial transcribed region where RNA synthesis can stall. In the third step, transcripts can be made as polymerase is released from these stalled positions with the assistance of an ATP-dependent process, likely phosphorylation of the polymerase C-terminal domain. After this third step, the promoter becomes cleared, allowing for the reinitiation of transcription.

The Full Text of this article is available as a PDF (336K).

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

• 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. [PubMed] [Google Scholar]
• Cadena DL, Dahmus ME. Messenger RNA synthesis in mammalian cells is catalyzed by the phosphorylated form of RNA polymerase II. J Biol Chem. 1987 Sep 15;262(26):12468–12474. [PubMed] [Google Scholar]
• Carey M, Leatherwood J, Ptashne M. A potent GAL4 derivative activates transcription at a distance in vitro. Science. 1990 Feb 9;247(4943):710–712. [PubMed] [Google Scholar]
• Carey M, Lin YS, Green MR, Ptashne M. A mechanism for synergistic activation of a mammalian gene by GAL4 derivatives. Nature. 1990 May 24;345(6273):361–364. [PubMed] [Google Scholar]
• Carpousis AJ, Gralla JD. Interaction of RNA polymerase with lacUV5 promoter DNA during mRNA initiation and elongation. Footprinting, methylation, and rifampicin-sensitivity changes accompanying transcription initiation. J Mol Biol. 1985 May 25;183(2):165–177. [PubMed] [Google Scholar]
• Conaway RC, Conaway JW. 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]
• 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] [Google Scholar]
• Drapkin R, Reinberg D. The multifunctional TFIIH complex and transcriptional control. Trends Biochem Sci. 1994 Nov;19(11):504–508. [PubMed] [Google Scholar]
• Ellinger T, Behnke D, Bujard H, Gralla JD. Stalling of Escherichia coli RNA polymerase in the +6 to +12 region in vivo is associated with tight binding to consensus promoter elements. J Mol Biol. 1994 Jun 17;239(4):455–465. [PubMed] [Google Scholar]
• Feaver WJ, Gileadi O, Li Y, Kornberg RD. CTD kinase associated with yeast RNA polymerase II initiation factor b. Cell. 1991 Dec 20;67(6):1223–1230. [PubMed] [Google Scholar]
• Feaver WJ, Svejstrup JQ, Henry NL, Kornberg RD. Relationship of CDK-activating kinase and RNA polymerase II CTD kinase TFIIH/TFIIK. Cell. 1994 Dec 16;79(6):1103–1109. [PubMed] [Google Scholar]
• 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] [Google Scholar]
• Goodrich JA, Tjian R. Transcription factors IIE and IIH and ATP hydrolysis direct promoter clearance by RNA polymerase II. Cell. 1994 Apr 8;77(1):145–156. [PubMed] [Google Scholar]
• Greenblatt J, Nodwell JR, Mason SW. Transcriptional antitermination. Nature. 1993 Jul 29;364(6436):401–406. [PubMed] [Google Scholar]
• Jacob GA, Luse SW, Luse DS. Abortive initiation is increased only for the weakest members of a set of down mutants of the adenovirus 2 major late promoter. J Biol Chem. 1991 Nov 25;266(33):22537–22544. [PubMed] [Google Scholar]
• Jacob GA, Kitzmiller JA, Luse DS. RNA polymerase II promoter strength in vitro may be reduced by defects at initiation or promoter clearance. J Biol Chem. 1994 Feb 4;269(5):3655–3663. [PubMed] [Google Scholar]
• Jiang Y, Gralla JD. Uncoupling of initiation and reinitiation rates during HeLa RNA polymerase II transcription in vitro. Mol Cell Biol. 1993 Aug;13(8):4572–4577. [PMC free article] [PubMed] [Google Scholar]
• Jiang Y, Gralla JD. RNA polymerase II phosphorylation: uncoupling from GAL4-VP16 directed open complex formation and transcription in a reconstituted system. Nucleic Acids Res. 1994 Nov 25;22(23):4958–4962. [PMC free article] [PubMed] [Google Scholar]
• Jiang Y, Gralla JD. Nucleotide requirements for activated RNA polymerase II open complex formation in vitro. J Biol Chem. 1995 Jan 20;270(3):1277–1281. [PubMed] [Google Scholar]
• Jiang Y, Smale ST, Gralla JD. A common ATP requirement for open complex formation and transcription at promoters containing initiator or TATA elements. J Biol Chem. 1993 Mar 25;268(9):6535–6540. [PubMed] [Google Scholar]
• Jiang Y, Triezenberg SJ, Gralla JD. Defective transcriptional activation by diverse VP16 mutants associated with a common inability to form open promoter complexes. J Biol Chem. 1994 Feb 25;269(8):5505–5508. [PubMed] [Google Scholar]
• Jiang Y, Yan M, Gralla JD. Abortive initiation and first bond formation at an activated adenovirus E4 promoter. J Biol Chem. 1995 Nov 10;270(45):27332–27338. [PubMed] [Google Scholar]
• 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] [Google Scholar]
• Kim YJ, Björklund S, Li Y, Sayre MH, Kornberg RD. A multiprotein mediator of transcriptional activation and its interaction with the C-terminal repeat domain of RNA polymerase II. Cell. 1994 May 20;77(4):599–608. [PubMed] [Google Scholar]
• Koleske AJ, Young RA. An RNA polymerase II holoenzyme responsive to activators. Nature. 1994 Mar 31;368(6470):466–469. [PubMed] [Google Scholar]
• Krumm A, Groudine M. Tumor suppression and transcription elongation: the dire consequences of changing partners. Science. 1995 Sep 8;269(5229):1400–1401. [PubMed] [Google Scholar]
• Krumm A, Hickey LB, Groudine M. Promoter-proximal pausing of RNA polymerase II defines a general rate-limiting step after transcription initiation. Genes Dev. 1995 Mar 1;9(5):559–572. [PubMed] [Google Scholar]
• Krumm A, Meulia T, Brunvand M, Groudine M. The block to transcriptional elongation within the human c-myc gene is determined in the promoter-proximal region. Genes Dev. 1992 Nov;6(11):2201–2213. [PubMed] [Google Scholar]
• 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] [Google Scholar]
• Laybourn PJ, Dahmus ME. Transcription-dependent structural changes in the C-terminal domain of mammalian RNA polymerase subunit IIa/o. J Biol Chem. 1989 Apr 25;264(12):6693–6698. [PubMed] [Google Scholar]
• Lee H, Kraus KW, Wolfner MF, Lis JT. DNA sequence requirements for generating paused polymerase at the start of hsp70. Genes Dev. 1992 Feb;6(2):284–295. [PubMed] [Google Scholar]
• Li Y, Kornberg RD. Interplay of positive and negative effectors in function of the C-terminal repeat domain of RNA polymerase II. Proc Natl Acad Sci U S A. 1994 Mar 15;91(6):2362–2366. [PMC free article] [PubMed] [Google Scholar]
• Liao SM, Zhang J, Jeffery DA, Koleske AJ, Thompson CM, Chao DM, Viljoen M, van Vuuren HJ, Young RA. A kinase-cyclin pair in the RNA polymerase II holoenzyme. Nature. 1995 Mar 9;374(6518):193–196. [PubMed] [Google Scholar]
• 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. [PMC free article] [PubMed] [Google Scholar]
• Lu H, Zawel L, Fisher L, Egly JM, Reinberg D. Human general transcription factor IIH phosphorylates the C-terminal domain of RNA polymerase II. Nature. 1992 Aug 20;358(6388):641–645. [PubMed] [Google Scholar]
• McClure WR. Mechanism and control of transcription initiation in prokaryotes. Annu Rev Biochem. 1985;54:171–204. [PubMed] [Google Scholar]
• O'Brien T, Hardin S, Greenleaf A, Lis JT. Phosphorylation of RNA polymerase II C-terminal domain and transcriptional elongation. Nature. 1994 Jul 7;370(6484):75–77. [PubMed] [Google Scholar]
• Parvin JD, Sharp PA. DNA topology and a minimal set of basal factors for transcription by RNA polymerase II. Cell. 1993 May 7;73(3):533–540. [PubMed] [Google Scholar]
• Payne JM, Laybourn PJ, Dahmus ME. The transition of RNA polymerase II from initiation to elongation is associated with phosphorylation of the carboxyl-terminal domain of subunit IIa. J Biol Chem. 1989 Nov 25;264(33):19621–19629. [PubMed] [Google Scholar]
• Roeder RG. The complexities of eukaryotic transcription initiation: regulation of preinitiation complex assembly. Trends Biochem Sci. 1991 Nov;16(11):402–408. [PubMed] [Google Scholar]
• Roy R, Adamczewski JP, Seroz T, Vermeulen W, Tassan JP, Schaeffer L, Nigg EA, Hoeijmakers JH, Egly JM. The MO15 cell cycle kinase is associated with the TFIIH transcription-DNA repair factor. Cell. 1994 Dec 16;79(6):1093–1101. [PubMed] [Google Scholar]
• Sawadogo M, Roeder RG. 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]
• Schaeffer L, Roy R, Humbert S, Moncollin V, Vermeulen W, Hoeijmakers JH, Chambon P, Egly JM. DNA repair helicase: a component of BTF2 (TFIIH) basic transcription factor. Science. 1993 Apr 2;260(5104):58–63. [PubMed] [Google Scholar]
• Serizawa H, Conaway JW, Conaway RC. Phosphorylation of C-terminal domain of RNA polymerase II is not required in basal transcription. Nature. 1993 May 27;363(6427):371–374. [PubMed] [Google Scholar]
• Serizawa H, Conaway RC, Conaway JW. 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. [PMC free article] [PubMed] [Google Scholar]
• Serizawa H, Conaway RC, Conaway JW. Multifunctional RNA polymerase II initiation factor delta from rat liver. Relationship between carboxyl-terminal domain kinase, ATPase, and DNA helicase activities. J Biol Chem. 1993 Aug 15;268(23):17300–17308. [PubMed] [Google Scholar]
• Serizawa H, Mäkelä TP, Conaway JW, Conaway RC, Weinberg RA, Young RA. Association of Cdk-activating kinase subunits with transcription factor TFIIH. Nature. 1995 Mar 16;374(6519):280–282. [PubMed] [Google Scholar]
• Shiekhattar R, Mermelstein F, Fisher RP, Drapkin R, Dynlacht B, Wessling HC, Morgan DO, Reinberg D. Cdk-activating kinase complex is a component of human transcription factor TFIIH. Nature. 1995 Mar 16;374(6519):283–287. [PubMed] [Google Scholar]
• Strobl LJ, Eick D. Hold back of RNA polymerase II at the transcription start site mediates down-regulation of c-myc in vivo. EMBO J. 1992 Sep;11(9):3307–3314. [PMC free article] [PubMed] [Google Scholar]
• Svejstrup JQ, Wang Z, Feaver WJ, Wu X, Bushnell DA, Donahue TF, Friedberg EC, Kornberg RD. Different forms of TFIIH for transcription and DNA repair: holo-TFIIH and a nucleotide excision repairosome. Cell. 1995 Jan 13;80(1):21–28. [PubMed] [Google Scholar]
• Tantin D, Carey M. A heteroduplex template circumvents the energetic requirement for ATP during activated transcription by RNA polymerase II. J Biol Chem. 1994 Jul 1;269(26):17397–17400. [PubMed] [Google Scholar]
• Thompson CM, Koleske AJ, Chao DM, Young RA. A multisubunit complex associated with the RNA polymerase II CTD and TATA-binding protein in yeast. Cell. 1993 Jul 2;73(7):1361–1375. [PubMed] [Google Scholar]
• Thompson NE, Aronson DB, Burgess RR. Purification of eukaryotic RNA polymerase II by immunoaffinity chromatography. Elution of active enzyme with protein stabilizing agents from a polyol-responsive monoclonal antibody. J Biol Chem. 1990 Apr 25;265(12):7069–7077. [PubMed] [Google Scholar]
• Thompson NE, Steinberg TH, Aronson DB, Burgess RR. Inhibition of in vivo and in vitro transcription by monoclonal antibodies prepared against wheat germ RNA polymerase II that react with the heptapeptide repeat of eukaryotic RNA polymerase II. J Biol Chem. 1989 Jul 5;264(19):11511–11520. [PubMed] [Google Scholar]
• Timmers HT. Transcription initiation by RNA polymerase II does not require hydrolysis of the beta-gamma phosphoanhydride bond of ATP. EMBO J. 1994 Jan 15;13(2):391–399. [PMC free article] [PubMed] [Google Scholar]
• Tyree CM, George CP, Lira-DeVito LM, Wampler SL, Dahmus ME, Zawel L, Kadonaga JT. 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. [PubMed] [Google Scholar]
• Usheva A, Maldonado E, Goldring A, Lu H, Houbavi C, Reinberg D, Aloni Y. Specific interaction between the nonphosphorylated form of RNA polymerase II and the TATA-binding protein. Cell. 1992 May 29;69(5):871–881. [PubMed] [Google Scholar]
• Van Dyke MW, Sawadogo M, Roeder RG. Stability of transcription complexes on class II genes. Mol Cell Biol. 1989 Jan;9(1):342–344. [PMC free article] [PubMed] [Google Scholar]
• Wang W, Carey M, Gralla JD. Polymerase II promoter activation: closed complex formation and ATP-driven start site opening. Science. 1992 Jan 24;255(5043):450–453. [PubMed] [Google Scholar]
• Wang W, Gralla JD, 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. [PubMed] [Google Scholar]
• Zawel L, Kumar KP, Reinberg D. Recycling of the general transcription factors during RNA polymerase II transcription. Genes Dev. 1995 Jun 15;9(12):1479–1490. [PubMed] [Google Scholar]

Page 2

Articles from this journal are generally available in PMC after a 6-month delay (embargo); however, the delay may vary at the discretion of the publisher.

Articles from Molecular and Cellular Biology are provided here courtesy of American Society for Microbiology (ASM)