Volume 271, Number 38, Issue of September 20, 1996 pp. 23352-23356
©1996 by The American Society for Biochemistry and Molecular Biology, Inc.

---

Promoter Escape by RNA Polymerase II
A ROLE FOR AN ATP COFACTOR IN SUPPRESSION OF ARREST BY POLYMERASE AT PROMOTER-PROXIMAL SITES

(Received for publication, July 15, 1996)

From the Program in Molecular and Cell Biology, Oklahoma Medical Research Foundation, Oklahoma City, Oklahoma 73104

It is well established that TFIIH-dependent transcription by RNA polymerase II requires a hydrolyzable ATP cofactor for synthesis of the first phosphodiester bond of nascent transcripts. Whether an ATP cofactor is also required after initiation for escape of RNA polymerase II from the promoter has, however, been controversial. We have now addressed this question directly by investigating the ability of RNA polymerase II transcription complexes containing short, ~5-8-nucleotide transcripts synthesized in the presence of limiting nucleotides to escape the promoter in the absence of an ATP cofactor in a basal transcription system reconstituted with purified RNA polymerase II and general initiation factors. Depletion of ATP had a profound effect on the ability of initiated complexes to progress into the elongation phase: whereas in the presence of ATP, the majority of transcription complexes could be chased away from the promoter-proximal region, most complexes deprived of ATP catalyzed synthesis of only a few phosphodiester bonds and then ceased elongation after synthesizing transcripts less than 10-14 nucleotides in length. A significant fraction of these transcripts could be extended following addition of ATP, indicating that they were contained in arrested, but potentially active elongation complexes. Like the ATP-requiring step in initiation, ATP-dependent suppression of arrest by RNA polymerase II at promoter-proximal sites is inhibited by adenosine 5-O-(thio)triphosphate. Transcription complexes containing transcripts longer than 9-10 nucleotides are insensitive to inhibition by ATPS, indicating that susceptibility to ATP-sensitive arrest is a property of very early elongation complexes. Taken together, our findings reveal a novel role for an ATP cofactor in transcription by RNA polymerase II.

CiteULike    Complore    Connotea    Del.icio.us    Digg    Reddit    Technorati    What's this?

This article has been cited by other articles:

				R. J. Sims III, R. Belotserkovskaya, and D. Reinberg Elongation by RNA polymerase II: the short and long of it Genes & Dev., October 15, 2004; 18(20): 2437 - 2468. [Abstract] [Full Text] [PDF]

				X. Wang, L. Spangler, and A. Dvir Promoter Escape by RNA Polymerase II. DOWNSTREAM PROMOTER DNA IS REQUIRED DURING MULTIPLE STEPS OF EARLY TRANSCRIPTION J. Biol. Chem., March 14, 2003; 278(12): 10250 - 10256. [Abstract] [Full Text] [PDF]

				M. Pal and D. S. Luse Strong Natural Pausing by RNA Polymerase II within 10 Bases of Transcription Start May Result in Repeated Slippage and Reextension of the Nascent RNA Mol. Cell. Biol., January 1, 2002; 22(1): 30 - 40. [Abstract] [Full Text] [PDF]

				M. Pal, D. McKean, and D. S. Luse Promoter Clearance by RNA Polymerase II Is an Extended, Multistep Process Strongly Affected by Sequence Mol. Cell. Biol., September 1, 2001; 21(17): 5815 - 5825. [Abstract] [Full Text] [PDF]

				L. Spangler, X. Wang, J. W. Conaway, R. C. Conaway, and A. Dvir TFIIH action in transcription initiation and promoter escape requires distinct regions of downstream promoter DNA PNAS, April 25, 2001; (2001) 101004498. [Abstract] [Full Text]

				J. W. Steinke, S. J. Kopytek, and D. O. Peterson Discrete promoter elements affect specific properties of RNA polymerase II transcription complexes Nucleic Acids Res., July 15, 2000; 28(14): 2726 - 2735. [Abstract] [Full Text] [PDF]

				T. Kim, R. H. Ebright, and D. Reinberg Mechanism of ATP-Dependent Promoter Melting by Transcription Factor IIH Science, May 26, 2000; 288(5470): 1418 - 1421. [Abstract] [Full Text]

				H. Tang, Y. Liu, L. Madabusi, and D. S. Gilmour Promoter-Proximal Pausing on the hsp70 Promoter in Drosophila melanogaster Depends on the Upstream Regulator Mol. Cell. Biol., April 1, 2000; 20(7): 2569 - 2580. [Abstract] [Full Text]

				J. Bradsher, F. Coin, and J.-M. Egly Distinct Roles for the Helicases of TFIIH in Transcript Initiation and Promoter Escape J. Biol. Chem., January 28, 2000; 275(4): 2532 - 2538. [Abstract] [Full Text] [PDF]

				Q. Yan, R. J. Moreland, J. W. Conaway, and R. C. Conaway Dual Roles for Transcription Factor IIF in Promoter Escape by RNA Polymerase II J. Biol. Chem., December 10, 1999; 274(50): 35668 - 35675. [Abstract] [Full Text] [PDF]

				M. Yan and J. D. Gralla The Use of ATP and Initiating Nucleotides during Postrecruitment Steps at the Activated Adenovirus E4 Promoter J. Biol. Chem., December 3, 1999; 274(49): 34819 - 34824. [Abstract] [Full Text] [PDF]

				R. J. Moreland, F. Tirode, Q. Yan, J. W. Conaway, J.-M. Egly, and R. C. Conaway A Role for the TFIIH XPB DNA Helicase in Promoter Escape by RNA Polymerase II J. Biol. Chem., August 6, 1999; 274(32): 22127 - 22130. [Abstract] [Full Text] [PDF]

				R. G. Keene and D. S. Luse Initially Transcribed Sequences Strongly Affect the Extent of Abortive Initiation by RNA Polymerase II J. Biol. Chem., April 23, 1999; 274(17): 11526 - 11534. [Abstract] [Full Text] [PDF]

				W. L. de Laat, N. G.J. Jaspers, and J. H.J. Hoeijmakers Molecular mechanism of nucleotide excision repair Genes & Dev., April 1, 1999; 13(7): 768 - 785. [Full Text]

				E. Kershnar, S.-Y. Wu, and C. M. Chiang Immunoaffinity Purification and Functional Characterization of Human Transcription Factor IIH and RNA Polymerase II from Clonal Cell Lines That Conditionally Express Epitope-tagged Subunits of the Multiprotein Complexes J. Biol. Chem., December 18, 1998; 273(51): 34444 - 34453. [Abstract] [Full Text] [PDF]

				K. P. Kumar, S. Akoulitchev, and D. Reinberg Promoter-proximal stalling results from the inability to recruit transcription factor IIH to the transcription complex and is a regulated event PNAS, August 18, 1998; 95(17): 9767 - 9772. [Abstract] [Full Text] [PDF]

				J. F. Kugel and J. A. Goodrich Promoter escape limits the rate of RNA polymerase II transcription and is enhanced by TFIIE, TFIIH, and ATP on negatively supercoiled DNA PNAS, August 4, 1998; 95(16): 9232 - 9237. [Abstract] [Full Text] [PDF]

				P. Tijerina and M. H. Sayre A Debilitating Mutation in Transcription Factor IIE with Differential Effects on Gene Expression in Yeast J. Biol. Chem., January 9, 1998; 273(2): 1107 - 1113. [Abstract] [Full Text] [PDF]

				S. A. Brown and R. E. Kingston Disruption of downstream chromatin directed by a transcriptional activator Genes & Dev., December 1, 1997; 11(23): 3116 - 3121. [Abstract] [Full Text] [PDF]

				A. Dvir, S. Tan, J. W. Conaway, and R. C. Conaway Promoter Escape by RNA Polymerase II. FORMATION OF AN ESCAPE-COMPETENT TRANSCRIPTIONAL INTERMEDIATE IS A PREREQUISITE FOR EXIT OF POLYMERASE FROM THE PROMOTER J. Biol. Chem., November 7, 1997; 272(45): 28175 - 28178. [Abstract] [Full Text] [PDF]

				A. Dvir, R. C. Conaway, and J. W. Conaway A role for TFIIH in controlling the activity of early RNA polymerase II elongation complexes PNAS, August 19, 1997; 94(17): 9006 - 9010. [Abstract] [Full Text] [PDF]

				S. M. Orlicky, P. T. Tran, M. H. Sayre, and A. M. Edwards Dissociable Rpb4-Rpb7 Subassembly of RNA Polymerase II Binds to Single-strand Nucleic Acid and Mediates a Post-recruitment Step in Transcription Initiation J. Biol. Chem., March 23, 2001; 276(13): 10097 - 10102. [Abstract] [Full Text] [PDF]

				J. F. Kugel and J. A. Goodrich A Kinetic Model for the Early Steps of RNA Synthesis by Human RNA Polymerase II J. Biol. Chem., December 15, 2000; 275(51): 40483 - 40491. [Abstract] [Full Text] [PDF]

				B. Sandrock and J.-M. Egly A Yeast Four-hybrid System Identifies Cdk-activating Kinase as a Regulator of the XPD Helicase, a Subunit of Transcription Factor IIH J. Biol. Chem., September 14, 2001; 276(38): 35328 - 35333. [Abstract] [Full Text] [PDF]

				L. Spangler, X. Wang, J. W. Conaway, R. C. Conaway, and A. Dvir TFIIH action in transcription initiation and promoter escape requires distinct regions of downstream promoter DNA PNAS, May 8, 2001; 98(10): 5544 - 5549. [Abstract] [Full Text] [PDF]