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. 1996 Dec;16(12):7031–7042. doi: 10.1128/mcb.16.12.7031

RNA polymerase III transcription from the human U6 and adenovirus type 2 VAI promoters has different requirements for human BRF, a subunit of human TFIIIB.

R Mital 1, R Kobayashi 1, N Hernandez 1
PMCID: PMC231706  PMID: 8943358

Abstract

Mammalian TFIIIB can be separated into two fractions required for transcription of the adenovirus type 2 VAI gene, which have been designated 0.38M-TFIIIB and 0.48M-TFIIIB. While 0.48M-TFIIIB has not been characterized, 0.38M-TFIIIB corresponds to a TBP-containing complex. We describe here the purification of this complex, which consists of TBP and a closely associated polypeptide of 88 kDa, and the isolation of a cDNA corresponding to the 88-kDa polypeptide. The predicted protein sequence reveals that the 88-kDa polypeptide corresponds to a human homolog of the Saccharomyces cerevisiae BRF protein, a subunit of yeast TFIIIB. Human BRF (hBRF) probably corresponds to TFIIIB90, a protein previously cloned by Wang and Roeder (Proc. Natl. Acad. Sci. USA 92:7026-7030, 1995), although its predicted amino acid sequence differs from that reported for TFIIIB90 over a stretch of 67 amino acids as a result of frameshifts. Immunodepletion of more than 90 to 95% of the hBRF present in a transcription extract severely debilitates transcription from the tRNA-type VAI promoter but does not affect transcription from the TATA box-containing human U6 promoter, suggesting that the 0.38M-TFIIIB complex, and perhaps hBRF as well, is not required for U6 transcription.

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Selected References

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  1. Berg J. M., Shi Y. The galvanization of biology: a growing appreciation for the roles of zinc. Science. 1996 Feb 23;271(5252):1081–1085. doi: 10.1126/science.271.5252.1081. [DOI] [PubMed] [Google Scholar]
  2. Brow D. A., Guthrie C. Transcription of a yeast U6 snRNA gene requires a polymerase III promoter element in a novel position. Genes Dev. 1990 Aug;4(8):1345–1356. doi: 10.1101/gad.4.8.1345. [DOI] [PubMed] [Google Scholar]
  3. Buratowski S., Zhou H. A suppressor of TBP mutations encodes an RNA polymerase III transcription factor with homology to TFIIB. Cell. 1992 Oct 16;71(2):221–230. doi: 10.1016/0092-8674(92)90351-c. [DOI] [PubMed] [Google Scholar]
  4. Burnol A. F., Margottin F., Huet J., Almouzni G., Prioleau M. N., Méchali M., Sentenac A. TFIIIC relieves repression of U6 snRNA transcription by chromatin. Nature. 1993 Apr 1;362(6419):475–477. doi: 10.1038/362475a0. [DOI] [PubMed] [Google Scholar]
  5. Burnol A. F., Margottin F., Schultz P., Marsolier M. C., Oudet P., Sentenac A. Basal promoter and enhancer element of yeast U6 snRNA gene. J Mol Biol. 1993 Oct 20;233(4):644–658. doi: 10.1006/jmbi.1993.1542. [DOI] [PubMed] [Google Scholar]
  6. Chiang C. M., Ge H., Wang Z., Hoffmann A., Roeder R. G. Unique TATA-binding protein-containing complexes and cofactors involved in transcription by RNA polymerases II and III. EMBO J. 1993 Jul;12(7):2749–2762. doi: 10.1002/j.1460-2075.1993.tb05936.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Colbert T., Hahn S. A yeast TFIIB-related factor involved in RNA polymerase III transcription. Genes Dev. 1992 Oct;6(10):1940–1949. doi: 10.1101/gad.6.10.1940. [DOI] [PubMed] [Google Scholar]
  8. Coleman J. E. Zinc proteins: enzymes, storage proteins, transcription factors, and replication proteins. Annu Rev Biochem. 1992;61:897–946. doi: 10.1146/annurev.bi.61.070192.004341. [DOI] [PubMed] [Google Scholar]
  9. Dean N., Berk A. J. Separation of TFIIIC into two functional components by sequence specific DNA affinity chromatography. Nucleic Acids Res. 1987 Dec 10;15(23):9895–9907. doi: 10.1093/nar/15.23.9895. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Eschenlauer J. B., Kaiser M. W., Gerlach V. L., Brow D. A. Architecture of a yeast U6 RNA gene promoter. Mol Cell Biol. 1993 May;13(5):3015–3026. doi: 10.1128/mcb.13.5.3015. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Frohman M. A. Rapid amplification of complementary DNA ends for generation of full-length complementary DNAs: thermal RACE. Methods Enzymol. 1993;218:340–356. doi: 10.1016/0076-6879(93)18026-9. [DOI] [PubMed] [Google Scholar]
  12. Gerlach V. L., Whitehall S. K., Geiduschek E. P., Brow D. A. TFIIIB placement on a yeast U6 RNA gene in vivo is directed primarily by TFIIIC rather than by sequence-specific DNA contacts. Mol Cell Biol. 1995 Mar;15(3):1455–1466. doi: 10.1128/mcb.15.3.1455. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Goomer R. S., Urso O., Kunkel G. R. A complex that contains proteins binding to the PSE and TATA sites in a human U6 small nuclear RNA promoter. Gene. 1994 Oct 21;148(2):269–275. doi: 10.1016/0378-1119(94)90698-x. [DOI] [PubMed] [Google Scholar]
  14. Henry R. W., Sadowski C. L., Kobayashi R., Hernandez N. A TBP-TAF complex required for transcription of human snRNA genes by RNA polymerase II and III. Nature. 1995 Apr 13;374(6523):653–656. doi: 10.1038/374653a0. [DOI] [PubMed] [Google Scholar]
  15. Hernandez N. TBP, a universal eukaryotic transcription factor? Genes Dev. 1993 Jul;7(7B):1291–1308. doi: 10.1101/gad.7.7b.1291. [DOI] [PubMed] [Google Scholar]
  16. Huet J., Conesa C., Manaud N., Chaussivert N., Sentenac A. Interactions between yeast TFIIIB components. Nucleic Acids Res. 1994 Aug 25;22(16):3433–3439. doi: 10.1093/nar/22.16.3433. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Joazeiro C. A., Kassavetis G. A., Geiduschek E. P. Alternative outcomes in assembly of promoter complexes: the roles of TBP and a flexible linker in placing TFIIIB on tRNA genes. Genes Dev. 1996 Mar 15;10(6):725–739. doi: 10.1101/gad.10.6.725. [DOI] [PubMed] [Google Scholar]
  18. Joazeiro C. A., Kassavetis G. A., Geiduschek E. P. Identical components of yeast transcription factor IIIB are required and sufficient for transcription of TATA box-containing and TATA-less genes. Mol Cell Biol. 1994 Apr;14(4):2798–2808. doi: 10.1128/mcb.14.4.2798. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Kassavetis G. A., Bartholomew B., Blanco J. A., Johnson T. E., Geiduschek E. P. Two essential components of the Saccharomyces cerevisiae transcription factor TFIIIB: transcription and DNA-binding properties. Proc Natl Acad Sci U S A. 1991 Aug 15;88(16):7308–7312. doi: 10.1073/pnas.88.16.7308. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Kassavetis G. A., Braun B. R., Nguyen L. H., Geiduschek E. P. S. cerevisiae TFIIIB is the transcription initiation factor proper of RNA polymerase III, while TFIIIA and TFIIIC are assembly factors. Cell. 1990 Jan 26;60(2):235–245. doi: 10.1016/0092-8674(90)90739-2. [DOI] [PubMed] [Google Scholar]
  21. Kassavetis G. A., Nguyen S. T., Kobayashi R., Kumar A., Geiduschek E. P., Pisano M. Cloning, expression, and function of TFC5, the gene encoding the B" component of the Saccharomyces cerevisiae RNA polymerase III transcription factor TFIIIB. Proc Natl Acad Sci U S A. 1995 Oct 10;92(21):9786–9790. doi: 10.1073/pnas.92.21.9786. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Khoo B., Brophy B., Jackson S. P. Conserved functional domains of the RNA polymerase III general transcription factor BRF. Genes Dev. 1994 Dec 1;8(23):2879–2890. doi: 10.1101/gad.8.23.2879. [DOI] [PubMed] [Google Scholar]
  23. Kovelman R., Roeder R. G. Purification and characterization of two forms of human transcription factor IIIC. J Biol Chem. 1992 Dec 5;267(34):24446–24456. [PubMed] [Google Scholar]
  24. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  25. Lai J. S., Cleary M. A., Herr W. A single amino acid exchange transfers VP16-induced positive control from the Oct-1 to the Oct-2 homeo domain. Genes Dev. 1992 Nov;6(11):2058–2065. doi: 10.1101/gad.6.11.2058. [DOI] [PubMed] [Google Scholar]
  26. Lobo S. M., Hernandez N. A 7 bp mutation converts a human RNA polymerase II snRNA promoter into an RNA polymerase III promoter. Cell. 1989 Jul 14;58(1):55–67. doi: 10.1016/0092-8674(89)90402-9. [DOI] [PubMed] [Google Scholar]
  27. Lobo S. M., Tanaka M., Sullivan M. L., Hernandez N. A TBP complex essential for transcription from TATA-less but not TATA-containing RNA polymerase III promoters is part of the TFIIIB fraction. Cell. 1992 Dec 11;71(6):1029–1040. doi: 10.1016/0092-8674(92)90397-u. [DOI] [PubMed] [Google Scholar]
  28. López-De-León A., Librizzi M., Puglia K., Willis I. M. PCF4 encodes an RNA polymerase III transcription factor with homology to TFIIB. Cell. 1992 Oct 16;71(2):211–220. doi: 10.1016/0092-8674(92)90350-l. [DOI] [PubMed] [Google Scholar]
  29. Margottin F., Dujardin G., Gérard M., Egly J. M., Huet J., Sentenac A. Participation of the TATA factor in transcription of the yeast U6 gene by RNA polymerase C. Science. 1991 Jan 25;251(4992):424–426. doi: 10.1126/science.1989075. [DOI] [PubMed] [Google Scholar]
  30. McBryant S. J., Meier E., Leresche A., Sharp S. J., Wolf V. J., Gottesfeld J. M. TATA-box DNA binding activity and subunit composition for RNA polymerase III transcription factor IIIB from Xenopus laevis. Mol Cell Biol. 1996 Sep;16(9):4639–4647. doi: 10.1128/mcb.16.9.4639. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Meyers R. E., Sharp P. A. TATA-binding protein and associated factors in polymerase II and polymerase III transcription. Mol Cell Biol. 1993 Dec;13(12):7953–7960. doi: 10.1128/mcb.13.12.7953. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Miller J., McLachlan A. D., Klug A. Repetitive zinc-binding domains in the protein transcription factor IIIA from Xenopus oocytes. EMBO J. 1985 Jun;4(6):1609–1614. doi: 10.1002/j.1460-2075.1985.tb03825.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Moenne A., Camier S., Anderson G., Margottin F., Beggs J., Sentenac A. The U6 gene of Saccharomyces cerevisiae is transcribed by RNA polymerase C (III) in vivo and in vitro. EMBO J. 1990 Jan;9(1):271–277. doi: 10.1002/j.1460-2075.1990.tb08105.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Murphy S., Yoon J. B., Gerster T., Roeder R. G. Oct-1 and Oct-2 potentiate functional interactions of a transcription factor with the proximal sequence element of small nuclear RNA genes. Mol Cell Biol. 1992 Jul;12(7):3247–3261. doi: 10.1128/mcb.12.7.3247. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Reddy R. Transcription of a U6 small nuclear RNA gene in vitro. Transcription of a mouse U6 small nuclear RNA gene in vitro by RNA polymerase III is dependent on transcription factor(s) different from transcription factors IIIA, IIIB, and IIIC. J Biol Chem. 1988 Nov 5;263(31):15980–15984. [PubMed] [Google Scholar]
  36. Reinberg D., Roeder R. G. Factors involved in specific transcription by mammalian RNA polymerase II. Transcription factor IIS stimulates elongation of RNA chains. J Biol Chem. 1987 Mar 5;262(7):3331–3337. [PubMed] [Google Scholar]
  37. Roberts S., Miller S. J., Lane W. S., Lee S., Hahn S. Cloning and functional characterization of the gene encoding the TFIIIB90 subunit of RNA polymerase III transcription factor TFIIIB. J Biol Chem. 1996 Jun 21;271(25):14903–14909. doi: 10.1074/jbc.271.25.14903. [DOI] [PubMed] [Google Scholar]
  38. Ruppert S. M., McCulloch V., Meyer M., Bautista C., Falkowski M., Stunnenberg H. G., Hernandez N. Monoclonal antibodies directed against the amino-terminal domain of human TBP cross-react with TBP from other species. Hybridoma. 1996 Feb;15(1):55–68. doi: 10.1089/hyb.1996.15.55. [DOI] [PubMed] [Google Scholar]
  39. Sadowski C. L., Henry R. W., Kobayashi R., Hernandez N. The SNAP45 subunit of the small nuclear RNA (snRNA) activating protein complex is required for RNA polymerase II and III snRNA gene transcription and interacts with the TATA box binding protein. Proc Natl Acad Sci U S A. 1996 Apr 30;93(9):4289–4293. doi: 10.1073/pnas.93.9.4289. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Sadowski C. L., Henry R. W., Lobo S. M., Hernandez N. Targeting TBP to a non-TATA box cis-regulatory element: a TBP-containing complex activates transcription from snRNA promoters through the PSE. Genes Dev. 1993 Aug;7(8):1535–1548. doi: 10.1101/gad.7.8.1535. [DOI] [PubMed] [Google Scholar]
  41. Segall J., Matsui T., Roeder R. G. Multiple factors are required for the accurate transcription of purified genes by RNA polymerase III. J Biol Chem. 1980 Dec 25;255(24):11986–11991. [PubMed] [Google Scholar]
  42. Shastry B. S., Ng S. Y., Roeder R. G. Multiple factors involved in the transcription of class III genes in Xenopus laevis. J Biol Chem. 1982 Nov 10;257(21):12979–12986. [PubMed] [Google Scholar]
  43. Skowronski J., Fanning T. G., Singer M. F. Unit-length line-1 transcripts in human teratocarcinoma cells. Mol Cell Biol. 1988 Apr;8(4):1385–1397. doi: 10.1128/mcb.8.4.1385. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Sparrow D. B., Wells J. R. Sequence of a cDNA encoding chicken high-mobility-group protein-2. Gene. 1992 May 15;114(2):289–290. doi: 10.1016/0378-1119(92)90590-l. [DOI] [PubMed] [Google Scholar]
  45. Taggart A. K., Fisher T. S., Pugh B. F. The TATA-binding protein and associated factors are components of pol III transcription factor TFIIIB. Cell. 1992 Dec 11;71(6):1015–1028. doi: 10.1016/0092-8674(92)90396-t. [DOI] [PubMed] [Google Scholar]
  46. Teichmann M., Seifart K. H. Physical separation of two different forms of human TFIIIB active in the transcription of the U6 or the VAI gene in vitro. EMBO J. 1995 Dec 1;14(23):5974–5983. doi: 10.1002/j.1460-2075.1995.tb00286.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Waldschmidt R., Wanandi I., Seifart K. H. Identification of transcription factors required for the expression of mammalian U6 genes in vitro. EMBO J. 1991 Sep;10(9):2595–2603. doi: 10.1002/j.1460-2075.1991.tb07801.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Wang Z., Roeder R. G. Structure and function of a human transcription factor TFIIIB subunit that is evolutionarily conserved and contains both TFIIB- and high-mobility-group protein 2-related domains. Proc Natl Acad Sci U S A. 1995 Jul 18;92(15):7026–7030. doi: 10.1073/pnas.92.15.7026. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. Whitehall S. K., Kassavetis G. A., Geiduschek E. P. The symmetry of the yeast U6 RNA gene's TATA box and the orientation of the TATA-binding protein in yeast TFIIIB. Genes Dev. 1995 Dec 1;9(23):2974–2985. doi: 10.1101/gad.9.23.2974. [DOI] [PubMed] [Google Scholar]
  50. Xiong Y., Hannon G. J., Zhang H., Casso D., Kobayashi R., Beach D. p21 is a universal inhibitor of cyclin kinases. Nature. 1993 Dec 16;366(6456):701–704. doi: 10.1038/366701a0. [DOI] [PubMed] [Google Scholar]
  51. Yoon J. B., Murphy S., Bai L., Wang Z., Roeder R. G. Proximal sequence element-binding transcription factor (PTF) is a multisubunit complex required for transcription of both RNA polymerase II- and RNA polymerase III-dependent small nuclear RNA genes. Mol Cell Biol. 1995 Apr;15(4):2019–2027. doi: 10.1128/mcb.15.4.2019. [DOI] [PMC free article] [PubMed] [Google Scholar]
  52. Yoon J. B., Roeder R. G. Cloning of two proximal sequence element-binding transcription factor subunits (gamma and delta) that are required for transcription of small nuclear RNA genes by RNA polymerases II and III and interact with the TATA-binding protein. Mol Cell Biol. 1996 Jan;16(1):1–9. doi: 10.1128/mcb.16.1.1. [DOI] [PMC free article] [PubMed] [Google Scholar]
  53. Yoshinaga S. K., Boulanger P. A., Berk A. J. Resolution of human transcription factor TFIIIC into two functional components. Proc Natl Acad Sci U S A. 1987 Jun;84(11):3585–3589. doi: 10.1073/pnas.84.11.3585. [DOI] [PMC free article] [PubMed] [Google Scholar]
  54. Yoshinaga S. K., L'Etoile N. D., Berk A. J. Purification and characterization of transcription factor IIIC2. J Biol Chem. 1989 Jun 25;264(18):10726–10731. [PubMed] [Google Scholar]

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