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. 1993 Feb;13(2):1296–1305. doi: 10.1128/mcb.13.2.1296

Transcription elongation in the human c-myc gene is governed by overall transcription initiation levels in Xenopus oocytes.

C A Spencer 1, M A Kilvert 1
PMCID: PMC359015  PMID: 8423795

Abstract

Both transcription initiation and transcription elongation contribute to the regulation of steady-state c-myc RNA levels. We have used the Xenopus oocyte transcription assay to study premature transcription termination which occurs in the first exon and intron of the human c-myc gene. Previous studies showed that after injection into Xenopus oocytes transcription from the c-myc P1 promoter resulted in read-through transcripts whereas transcription from the stronger P2 promoter resulted in a combination of prematurely terminated and read-through transcripts. We now demonstrate that this promoter-specific processivity results from the overall amount of RNA polymerase II transcription occurring from either promoter. Parameters that reduce the amount of transcription from P1 or P2, such as decreased concentration of template injected or decreased incubation time, result in a reduction in the ratio of terminated to read-through c-myc transcripts. Conversely, when transcription levels are increased by higher concentrations of injected template, increased incubation time, or coinjection with competing template, the ratio of terminated to read-through transcripts increases. We hypothesize that an RNA polymerase II processivity function is depleted above a threshold level of transcription initiation, resulting in high levels of premature transcription termination. These findings account for the promoter-specific effects on transcription elongation previously seen in this assay system and suggest a mechanism whereby limiting transcription elongation factors may contribute to transcription regulation in other eukaryotic cells.

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Selected References

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

  1. Bengal E., Flores O., Krauskopf A., Reinberg D., Aloni Y. Role of the mammalian transcription factors IIF, IIS, and IIX during elongation by RNA polymerase II. Mol Cell Biol. 1991 Mar;11(3):1195–1206. doi: 10.1128/mcb.11.3.1195. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bentley D. L., Groudine M. A block to elongation is largely responsible for decreased transcription of c-myc in differentiated HL60 cells. Nature. 1986 Jun 12;321(6071):702–706. doi: 10.1038/321702a0. [DOI] [PubMed] [Google Scholar]
  3. Bentley D. L., Groudine M. Sequence requirements for premature termination of transcription in the human c-myc gene. Cell. 1988 Apr 22;53(2):245–256. doi: 10.1016/0092-8674(88)90386-8. [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. Burton Z. F., Killeen M., Sopta M., Ortolan L. G., Greenblatt J. RAP30/74: a general initiation factor that binds to RNA polymerase II. Mol Cell Biol. 1988 Apr;8(4):1602–1613. doi: 10.1128/mcb.8.4.1602. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Chafin D. R., Claussen T. J., Price D. H. Identification and purification of a yeast protein that affects elongation by RNA polymerase II. J Biol Chem. 1991 May 15;266(14):9256–9262. [PubMed] [Google Scholar]
  7. Challoner P. B., Moss S. B., Groudine M. Expression of replication-dependent histone genes in avian spermatids involves an alternate pathway of mRNA 3'-end formation. Mol Cell Biol. 1989 Mar;9(3):902–913. doi: 10.1128/mcb.9.3.902. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Chen Z., Harless M. L., Wright D. A., Kellems R. E. Identification and characterization of transcriptional arrest sites in exon 1 of the human adenosine deaminase gene. Mol Cell Biol. 1990 Sep;10(9):4555–4564. doi: 10.1128/mcb.10.9.4555. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Chen Z., Innis J. W., Sun M. H., Wright D. A., Kellems R. E. Sequence requirements for transcriptional arrest in exon 1 of the human adenosine deaminase gene. Mol Cell Biol. 1991 Dec;11(12):6248–6256. doi: 10.1128/mcb.11.12.6248. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Chinsky J. M., Maa M. C., Ramamurthy V., Kellems R. E. Adenosine deaminase gene expression. Tissue-dependent regulation of transcriptional elongation. J Biol Chem. 1989 Aug 25;264(24):14561–14565. [PubMed] [Google Scholar]
  11. Choder M. A general topoisomerase I-dependent transcriptional repression in the stationary phase in yeast. Genes Dev. 1991 Dec;5(12A):2315–2326. doi: 10.1101/gad.5.12a.2315. [DOI] [PubMed] [Google Scholar]
  12. Fort P., Rech J., Vie A., Piechaczyk M., Bonnieu A., Jeanteur P., Blanchard J. M. Regulation of c-fos gene expression in hamster fibroblasts: initiation and elongation of transcription and mRNA degradation. Nucleic Acids Res. 1987 Jul 24;15(14):5657–5667. doi: 10.1093/nar/15.14.5657. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Grayhack E. J., Yang X. J., Lau L. F., Roberts J. W. Phage lambda gene Q antiterminator recognizes RNA polymerase near the promoter and accelerates it through a pause site. Cell. 1985 Aug;42(1):259–269. doi: 10.1016/s0092-8674(85)80121-5. [DOI] [PubMed] [Google Scholar]
  14. Haley J. D., Waterfield M. D. Contributory effects of de novo transcription and premature transcript termination in the regulation of human epidermal growth factor receptor proto-oncogene RNA synthesis. J Biol Chem. 1991 Jan 25;266(3):1746–1753. [PubMed] [Google Scholar]
  15. Hall D. J. Regulation of c-myc transcription in vitro: dependence on the guanine-rich promoter element ME1a1. Oncogene. 1990 Jan;5(1):47–54. [PubMed] [Google Scholar]
  16. Harland R. M., Weintraub H., McKnight S. L. Transcription of DNA injected into Xenopus oocytes is influenced by template topology. Nature. 1983 Mar 3;302(5903):38–43. doi: 10.1038/302038a0. [DOI] [PubMed] [Google Scholar]
  17. Hernandez N., Lucito R. Elements required for transcription initiation of the human U2 snRNA gene coincide with elements required for snRNA 3' end formation. EMBO J. 1988 Oct;7(10):3125–3134. doi: 10.1002/j.1460-2075.1988.tb03179.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Hernandez N., Weiner A. M. Formation of the 3' end of U1 snRNA requires compatible snRNA promoter elements. Cell. 1986 Oct 24;47(2):249–258. doi: 10.1016/0092-8674(86)90447-2. [DOI] [PubMed] [Google Scholar]
  19. Izban M. G., Luse D. S. Factor-stimulated RNA polymerase II transcribes at physiological elongation rates on naked DNA but very poorly on chromatin templates. J Biol Chem. 1992 Jul 5;267(19):13647–13655. [PubMed] [Google Scholar]
  20. Izban M. G., Luse D. S. The RNA polymerase II ternary complex cleaves the nascent transcript in a 3'----5' direction in the presence of elongation factor SII. Genes Dev. 1992 Jul;6(7):1342–1356. doi: 10.1101/gad.6.7.1342. [DOI] [PubMed] [Google Scholar]
  21. Kao S. Y., Calman A. F., Luciw P. A., Peterlin B. M. Anti-termination of transcription within the long terminal repeat of HIV-1 by tat gene product. Nature. 1987 Dec 3;330(6147):489–493. doi: 10.1038/330489a0. [DOI] [PubMed] [Google Scholar]
  22. Kerppola T. K., Kane C. M. Intrinsic sites of transcription termination and pausing in the c-myc gene. Mol Cell Biol. 1988 Oct;8(10):4389–4394. doi: 10.1128/mcb.8.10.4389. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Kerppola T. K., Kane C. M. RNA polymerase: regulation of transcript elongation and termination. FASEB J. 1991 Oct;5(13):2833–2842. doi: 10.1096/fasebj.5.13.1916107. [DOI] [PubMed] [Google Scholar]
  24. Kessler M., Ben-Asher E., Aloni Y. Elements modulating the block of transcription elongation at the adenovirus 2 attenuation site. J Biol Chem. 1989 Jun 15;264(17):9785–9790. [PubMed] [Google Scholar]
  25. Krystal G., Birrer M., Way J., Nau M., Sausville E., Thompson C., Minna J., Battey J. Multiple mechanisms for transcriptional regulation of the myc gene family in small-cell lung cancer. Mol Cell Biol. 1988 Aug;8(8):3373–3381. doi: 10.1128/mcb.8.8.3373. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Laspia M. F., Rice A. P., Mathews M. B. HIV-1 Tat protein increases transcriptional initiation and stabilizes elongation. Cell. 1989 Oct 20;59(2):283–292. doi: 10.1016/0092-8674(89)90290-0. [DOI] [PubMed] [Google Scholar]
  27. Lee H., Kraus K. W., Wolfner M. F., Lis J. T. DNA sequence requirements for generating paused polymerase at the start of hsp70. Genes Dev. 1992 Feb;6(2):284–295. doi: 10.1101/gad.6.2.284. [DOI] [PubMed] [Google Scholar]
  28. Lindsten T., June C. H., Thompson C. B. Multiple mechanisms regulate c-myc gene expression during normal T cell activation. EMBO J. 1988 Sep;7(9):2787–2794. doi: 10.1002/j.1460-2075.1988.tb03133.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Lois R., Freeman L., Villeponteau B., Martinson H. G. Active beta-globin gene transcription occurs in methylated, DNase I-resistant chromatin of nonerythroid chicken cells. Mol Cell Biol. 1990 Jan;10(1):16–27. doi: 10.1128/mcb.10.1.16. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Marshall N. F., Price D. H. Control of formation of two distinct classes of RNA polymerase II elongation complexes. Mol Cell Biol. 1992 May;12(5):2078–2090. doi: 10.1128/mcb.12.5.2078. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Mason S. W., Greenblatt J. Assembly of transcription elongation complexes containing the N protein of phage lambda and the Escherichia coli elongation factors NusA, NusB, NusG, and S10. Genes Dev. 1991 Aug;5(8):1504–1512. doi: 10.1101/gad.5.8.1504. [DOI] [PubMed] [Google Scholar]
  32. Meulia T., Krumm A., Spencer C., Groudine M. Sequences in the human c-myc P2 promoter affect the elongation and premature termination of transcripts initiated from the upstream P1 promoter. Mol Cell Biol. 1992 Oct;12(10):4590–4600. doi: 10.1128/mcb.12.10.4590. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Middleton K. M., Morgan G. T. Premature termination of transcription can be induced on an injected alpha-tubulin gene in Xenopus oocytes. Mol Cell Biol. 1990 Feb;10(2):727–735. doi: 10.1128/mcb.10.2.727. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Miller H., Asselin C., Dufort D., Yang J. Q., Gupta K., Marcu K. B., Nepveu A. A cis-acting element in the promoter region of the murine c-myc gene is necessary for transcriptional block. Mol Cell Biol. 1989 Dec;9(12):5340–5349. doi: 10.1128/mcb.9.12.5340. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Morrow M. A., Lee G., Gillis S., Yancopoulos G. D., Alt F. W. Interleukin-7 induces N-myc and c-myc expression in normal precursor B lymphocytes. Genes Dev. 1992 Jan;6(1):61–70. doi: 10.1101/gad.6.1.61. [DOI] [PubMed] [Google Scholar]
  36. Nepveu A., Marcu K. B. Intragenic pausing and anti-sense transcription within the murine c-myc locus. EMBO J. 1986 Nov;5(11):2859–2865. doi: 10.1002/j.1460-2075.1986.tb04580.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Neuman de Vegvar H. E., Dahlberg J. E. Initiation and termination of human U1 RNA transcription requires the concerted action of multiple flanking elements. Nucleic Acids Res. 1989 Nov 25;17(22):9305–9318. doi: 10.1093/nar/17.22.9305. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Price D. H., Sluder A. E., Greenleaf A. L. Dynamic interaction between a Drosophila transcription factor and RNA polymerase II. Mol Cell Biol. 1989 Apr;9(4):1465–1475. doi: 10.1128/mcb.9.4.1465. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Ramamurthy V., Maa M. C., Harless M. L., Wright D. A., Kellems R. E. Sequence requirements for transcriptional arrest in exon 1 of the murine adenosine deaminase gene. Mol Cell Biol. 1990 Apr;10(4):1484–1491. doi: 10.1128/mcb.10.4.1484. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Reines D., Chamberlin M. J., Kane C. M. Transcription elongation factor SII (TFIIS) enables RNA polymerase II to elongate through a block to transcription in a human gene in vitro. J Biol Chem. 1989 Jun 25;264(18):10799–10809. [PubMed] [Google Scholar]
  41. Reines D. Elongation factor-dependent transcript shortening by template-engaged RNA polymerase II. J Biol Chem. 1992 Feb 25;267(6):3795–3800. [PMC free article] [PubMed] [Google Scholar]
  42. Resnekov O., Kessler M., Aloni Y. RNA secondary structure is an integral part of the in vitro mechanism of attenuation in simian virus 40. J Biol Chem. 1989 Jun 15;264(17):9953–9959. [PubMed] [Google Scholar]
  43. Roberts J. W. Phage lambda and the regulation of transcription termination. Cell. 1988 Jan 15;52(1):5–6. doi: 10.1016/0092-8674(88)90523-5. [DOI] [PubMed] [Google Scholar]
  44. Roberts S., Bentley D. L. Distinct modes of transcription read through or terminate at the c-myc attenuator. EMBO J. 1992 Mar;11(3):1085–1093. doi: 10.1002/j.1460-2075.1992.tb05147.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Rougvie A. E., Lis J. T. The RNA polymerase II molecule at the 5' end of the uninduced hsp70 gene of D. melanogaster is transcriptionally engaged. Cell. 1988 Sep 9;54(6):795–804. doi: 10.1016/s0092-8674(88)91087-2. [DOI] [PubMed] [Google Scholar]
  46. Selby M. J., Bain E. S., Luciw P. A., Peterlin B. M. Structure, sequence, and position of the stem-loop in tar determine transcriptional elongation by tat through the HIV-1 long terminal repeat. Genes Dev. 1989 Apr;3(4):547–558. doi: 10.1101/gad.3.4.547. [DOI] [PubMed] [Google Scholar]
  47. Sharp P. A., Marciniak R. A. HIV TAR: an RNA enhancer? Cell. 1989 Oct 20;59(2):229–230. doi: 10.1016/0092-8674(89)90279-1. [DOI] [PubMed] [Google Scholar]
  48. Shermoen A. W., O'Farrell P. H. Progression of the cell cycle through mitosis leads to abortion of nascent transcripts. Cell. 1991 Oct 18;67(2):303–310. doi: 10.1016/0092-8674(91)90182-x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. Skarnes W. C., Tessier D. C., Acheson N. H. RNA polymerases stall and/or prematurely terminate nearby both early and late promoters on polyomavirus DNA. J Mol Biol. 1988 Sep 5;203(1):153–171. doi: 10.1016/0022-2836(88)90099-x. [DOI] [PubMed] [Google Scholar]
  50. Sluder A. E., Greenleaf A. L., Price D. H. Properties of a Drosophila RNA polymerase II elongation factor. J Biol Chem. 1989 May 25;264(15):8963–8969. [PubMed] [Google Scholar]
  51. Spencer C. A., Groudine M. Transcription elongation and eukaryotic gene regulation. Oncogene. 1990 Jun;5(6):777–785. [PubMed] [Google Scholar]
  52. Spencer C. A., LeStrange R. C., Novak U., Hayward W. S., Groudine M. The block to transcription elongation is promoter dependent in normal and Burkitt's lymphoma c-myc alleles. Genes Dev. 1990 Jan;4(1):75–88. doi: 10.1101/gad.4.1.75. [DOI] [PubMed] [Google Scholar]
  53. Walmsley M. E., Patient R. K. Highly efficient beta globin transcription in the absence of both a viral enhancer and erythroid factors. Development. 1987 Dec;101(4):815–827. doi: 10.1242/dev.101.4.815. [DOI] [PubMed] [Google Scholar]
  54. Watson R. J. A transcriptional arrest mechanism involved in controlling constitutive levels of mouse c-myb mRNA. Oncogene. 1988 Mar;2(3):267–272. [PubMed] [Google Scholar]
  55. Wiest D. K., Hawley D. K. In vitro analysis of a transcription termination site for RNA polymerase II. Mol Cell Biol. 1990 Nov;10(11):5782–5795. doi: 10.1128/mcb.10.11.5782. [DOI] [PMC free article] [PubMed] [Google Scholar]
  56. Wright S., Mirels L. F., Calayag M. C., Bishop J. M. Premature termination of transcription from the P1 promoter of the mouse c-myc gene. Proc Natl Acad Sci U S A. 1991 Dec 15;88(24):11383–11387. doi: 10.1073/pnas.88.24.11383. [DOI] [PMC free article] [PubMed] [Google Scholar]

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