RNAポリメラーゼの機能制御による転写包括制御の全体像

2004 Fiscal Year Annual Research Report

• Project/Area Number: 15370079
• Research Institution: Nippon Institute for Biological Science
• Principal Investigator: 石浜 明 (財団法人)日本生物科学研究所, 研究部, 主任研究員 (80019869)
• Co-Investigator(Kenkyū-buntansha): 本田 文江 (財団法人)日本生物科学研究所, 研究部, 研究員 (80343747) ; 岩田 晃 (財団法人)日本生物科学研究所, 研究部, 主任研究員 (70193745)
• Keywords: RNAポリメラーゼ / 転写因子 / シグマ因子 / プロモーター / 転写調節 / 大腸菌 / SELEX

Research Abstract

大腸菌転写装置(RNAポリメラーゼ>は、7種類のシグア因子、約300種類の転写因子との相互作用で、転写対象遺伝子選択特性を変化させ、その結果、ゲノム約4000遺伝子の転写パターンが変化する。転写装置の遺伝子間での分配制御の分子機構を解明する目的で、大腸菌各種転写因子に関して、支配下遺伝子群と各遺伝子の転写因子結合シグナルを同定し、転写因子による転写調節様式を解析し、結果を総合して転写装置の機能制御によるゲノム転写パーンの包括制御の全体像を解明することを目指した研究を実施した。以下は、成果の概要である。
1)全7種類シグマ因子、200種類の転写因子の精製
全シグマ因子、200種類の転写因子の発現系を構築し精製純化した。
2)全シグマ因子、転写因子50種類の支配下遺伝子の同定
断片化ゲノムから転写因子が結合するDNA断片を単離し、塩基配列から転写因子の結合部位を同定する新SELEX法を確立し、機能未知転写因子を中心に約50種類の転写因子の支配下遺伝子を同定した。
3)全シグマ因子、転写因子50種類の結合DNAシグナルの同定
SELEXで同定した転写因子支配下遺伝子上の転写因子結合部位をDNAフットプリント法及びS1マッピング法で同定した。
4)全シグマ因子、転写因子50種類の支配下プロモーター群の転写調節(in vitro)
転写因子支配下プロモーター、純化転写因子と純化RNAポリメラーゼを用いた試験管内転写系で利用して、試験転写因子による転写調節様式を解析した。
5)全シグマ因子、転写因子50種類の支配下プロモーター群の転写調節(in vivo解析)
新たに開発した、二種類の蛍光蛋白をレポーターとしてプロモーター活性を測定するTFPベクターに、SELEXで同定したプロモーター群を挿入し、生体内における、試験転写因子による転写調節様式を解析した。

Research Products

• [Journal Article] The fission yeast protein, Ker1p, is an ortholog of RNA polymerase I subunit A14 in Saccharomyces cerevisiae, and is required for stable association of Rrms and RPA21 in Pol I. (2005)
  • Author(s): Imazawa, Y., Hisatake, K., Mitsuzawa, H., Matsumoto, M., Tsukui, T., Nakagawa, K., Nakadai, T., Shimada, M., Ishihama, A., Nogi, Y
  • Journal Title: J.Biol.Chem. 280(12) Pages: 11467-11474
• [Journal Article] Glyceraldehyde-3-phosphate dehydrogenase associates with RNA polymerase II and interacts with its Rpb7 subunit. (2005)
  • Author(s): Mitsuzawa, H., Kimura, M., Kanda, E., Ishihama, A
  • Journal Title: FEBS Letters 579 Pages: 48-52
• [Journal Article] uvrA and pric genes involved in DNA repair and replication are negatively regulated by ArcA in Escherichia coli K12. (2005)
  • Author(s): Ogasawara, H., Teramoto, J., Hirao, K., Yamamoto, K., Ishihama, A., Utsumi, R.
  • Journal Title: J.Bactereiol. (in press)
• [Journal Article] Systematic search for the Cra-binding promoters using genomic SELEX. (2005)
  • Author(s): Shimada, T., Fujita, N., Maeda, M., Ishihama.A.
  • Journal Title: Genes Cells (in press) Pages: 215-227
• [Journal Article] Transcriptional response of Escherichia coil to external copper. (2005)
  • Author(s): Yamamoto, K., Ishihama A.
  • Journal Title: Mol.Microbiol. 55(1) Pages: 215-227
• [Journal Article] Cross-talks in signal transduction network among all two-component systems in Escherichia coli. (2005)
  • Author(s): Yamamoto, K., Hirano, K., Ohshima, T., Aiba, H., Utsumi, R., Ishihama.A.
  • Journal Title: J.Biol.Chem. 280(2) Pages: 1448-1456
• [Journal Article] Fundamental structural units of the Escherichia coli nucleoid revealed by atomic force microscopy. (2004)
  • Author(s): Kim, J., Yoshimura, S.H., Hizume, K., Ohniwa, R.L., Ishihama, A., Takeyasu, K.
  • Journal Title: Nucleic Acids Res. 32(6) Pages: 1982-1992
  • Description: 「研究成果報告書概要(欧文)」より
• [Journal Article] RNA polymerase II transcription apparatus in Schizosaccharomyces pombe. (2004)
  • Author(s): Mitsuzawa, H., Ishihama, A.
  • Journal Title: Curr.Genet. 44(6) Pages: 287-294
  • Description: 「研究成果報告書概要(欧文)」より
• [Journal Article] Modes of action of the TyrR protein, which result in repression and activation of the tyrP promoter of Escherichia coli. (2004)
  • Author(s): Yang, J., Hwang, S., Camakaris, H., Irawaty, W., Ishihama, A., Pittard, J.
  • Journal Title: Mol.Microbiol. 52(1) Pages: 243-256
  • Description: 「研究成果報告書概要(欧文)」より
• [Journal Article] Signal transduction cascade between EvgA/EvgS and PhoP/PhoQ two-component systems of Escherichia coli. (2004)
  • Author(s): Eguchi, Y., Okada, T., Minagawa, S., Oshima, T., Mori, H., Yamamoto, K., Ishihama A., Utsumi, R
  • Journal Title: J.Bacteriol. 186(10) Pages: 3006-3014
• [Journal Article] High level activity of (2',3'9-oligoadenylate synthetase in dog serum. (2004)
  • Author(s): Iwata, A., Yamamoto, A., Fujino, M., Sato, I., Hosokawa-Kanai, T., Tuchiya, K., Ishihamra A., Sokawa, Y
  • Journal Title: J.Vet.Med.Sci. 66(6) Pages: 721-724
• [Journal Article] A regulatory trade-off as a source of strain variation in the species Escherichia coli. (2004)
  • Author(s): King, T., Ishihama, A., Kori, A., Ferrenci, T
  • Journal Title: J.Bacteriol. 186(17) Pages: 5614-5620
• [Journal Article] Structure-Function Relationships of Influenza Virus RNA Plymerase : Primer-binding Site on PB1 Subunit. (2004)
  • Author(s): Kolpashchikov, D., Honda, A., Ishihama, A
  • Journal Title: Biochemistry 43(19) Pages: 5882-5887
• [Journal Article] Negative regulation of the DNA repair gene (ung) epression by CpxR/cxA two-component system in Escherichia coli and induction of mutations by increased expression of CpxR. (2004)
  • Author(s): Ogasawara, H., Teramoto, J., Yamamoto, K., Ishihama A., Utsumi, R
  • Journal Title: J.Bacteriol. 186(24) Pages: 8317-8325
• [Journal Article] Quantitative analysis of Escherichia coli promoter activity : Stationary-phase gene-associated promoters (2004)
  • Author(s): Shimada, T., Makinoshima, H., Ogawa, N., Maeda, M., Ishihama, A.
  • Journal Title: J.Bacteriol. 186(21) Pages: 7112-7122
• [Journal Article] Genome architecture studied by nano-scale imaging : Analyses among bacterial phyla and their implications to eukaryotic genome folding. (2004)
  • Author(s): Takeyasu, K., Kim, J., Ohniwa, R.L., Kobori, T., Inose, Y., Morikawa, K., Ohta, T., Ishihama, A., Yoshimura, S.H
  • Journal Title: Cytogenet.Genome Res. 107(1-2) Pages: 38-48
• [Journal Article] Identification and characterization of antibacterial agents that inhibit the essential histidine kinase, YycG, from Baccilus subtilis and Staphylococcus aureus. (2004)
  • Author(s): Watanabe, T., Hashimoto, Y., Yamamoto, K., Hirao, K., Ishiahma, A., Fukamizo, T., Hino, M., Utsumi, R
  • Journal Title: J.Antibiot. 56(12) Pages: 1045-1052
• [Journal Article] A unifying model for the role of polyamines in bacterial cell growth : the polyamine modulon. (2004)
  • Author(s): Yoshida, M., Kashiwagi, K., Shigemasa, A., Shigemasa, A., Taniguchi, S., Yamamoto, K., Makinoshima, H., Ishihama.A., Igarashi, K
  • Journal Title: J.Biol.Chem. 279(44) Pages: 46008-46013
• [Journal Article] Mapping sigma 54-RNA polymerase interactions at the -24 consensus promoter element. (2003)
  • Author(s): Burrows, P.C., Severinov, K., Ishihama, A., Buck, M., Wigneshweraraj, S.R.
  • Journal Title: J.Biol.Chem. 278(32) Pages: 29728-29743
  • Description: 「研究成果報告書概要(欧文)」より
• [Journal Article] Identification and molecular characterization of Mg(II)-regulon of Escherichia coli. (2003)
  • Author(s): Minagawa, S., Ogasawara, H., Kato, A., Yamamoto, K., Eguchi, T., Oshima, T., Mori, H., Ishihama, A., Utsumi, R.
  • Journal Title: J.Bacteriol. 185(13) Pages: 3696-3702
  • Description: 「研究成果報告書概要(欧文)」より
• [Journal Article] Rpb7 subunit of RNA polymerase II interacts with an RNA-binding protein involved in processing of transcripts. (2003)
  • Author(s): Mitsuzawa, H., Kanda, E., Ishihama, A.
  • Journal Title: Nucleic Acids Res. 31(16) Pages: 4696-4701
  • Description: 「研究成果報告書概要(欧文)」より
• [Journal Article] Role of a nonnatural b-C-nucleotide unit in DNA as template for DNA and RNA syntheses and as substrates for nucleolytic digestion. (2003)
  • Author(s): Aketani, S., Tanaka, Yamamoto, K., Ishihama, A., Cao, H., Tengenji, A., Shionoya, M.
  • Journal Title: Eur.J.Pharm.Sci. 20 Pages: 43-51
• [Journal Article] Transcription regulation of the drug efflux genes by EvgAS, a two-component system in Escherichia coli. (2003)
  • Author(s): Eguchi, Y., Oshima, T., Mori, H., Aono, R., Yamamoto, K., Ishihama.A.Utsumi, R.:
  • Journal Title: Microbiology 149(10) Pages: 2819-2828
• [Journal Article] In Vitro Properties of RpoS (σ^s) mutants of escherichia coli with postulated N-Terminal subregion 1.1 or C-terminal region 4 deleted. (2003)
  • Author(s): Gowrishankar, J., Yamamoto, K., Subbarayan, P.R., Ishihama.A.
  • Journal Title: J.Bacteriol. 185(8) Pages: 2673-2679
• [Journal Article] Involvement of region 4 of the sigma 70 subunit of RNA polymerase in transcription activation of the lux operon during quorum sensing. (2003)
  • Author(s): Johnson, D.C., Ishihama, A., Stevens, A.M
  • Journal Title: FEMS Microbiol. Lett. 228(2) Pages: 193-201
• [Journal Article] Recruitment of σ^<54>-RNA polymerase to the Pu promoter of Pseudomonas putida through integration host factor-mediated positioning switch of subunit carboxy-terminal domain on an UP-like element. (2003)
  • Author(s): Macchi, R., Montesissa, L., Murakami, K., Ishihama.A., de Lorenzo, V., Bertoni, G.:
  • Journal Title: J.Biol.Chem. 278(30) Pages: 27695-27702
• [Journal Article] Growth phase-coupled alterations in cell structure and function of Escherichia coli. (2003)
  • Author(s): Makinoshima, H., Aizawa, S., Hayashi, H., Miki, T., Nishimura, A., Ishihama, A
  • Journal Title: J.Bacteriol. 185(4) Pages: 1338-1345
• [Journal Article] The fission yeast RPA51 is a functional homolog of the budding yeast A49 subunit of RNA polymerase I and required for maximizing transcription of ribosomal RNA. (2003)
  • Author(s): Nakagawa, K., Hisatake, K., Imazawa, Y., Ishiguro, A., Matsumoto, M., Pape, L., Ishihama, A., Nogi, Y
  • Journal Title: Genes Genet.Syst. 78 Pages: 199-209
• [Journal Article] Identification and characterization of antibacterial agents that inhibit the essential histidine kinase, YycG, from Baccilus subtilis and Staphylococcus aureus. (2003)
  • Author(s): Watanabe, T., Hashimoto, Y., Yamamoto, K., Hirao, K., Ishiahma, A., Fukamizo, T., Hino, M., Utsumi, R.:
  • Journal Title: J.Antibiot. 56(12) Pages: 1045-1052
• [Journal Article] Two different modes of transcription repression of the Escherichia coli acetate operon by IcIR. (2003)
  • Author(s): Yamamoto, K., Ishihama, A.
  • Journal Title: Mol.Microbiol. 47(1) Pages: 183-194
• [Book] International Congress Series : Options for Control of Influenza Virus, V (Y.Kawaoka, ed.) (2004)
  • Author(s): Honda, A., Ishihama, A.
  • Total Pages: 175
  • Publisher: Elsevier B.V., Amsterdam
• [Book] The Molecular Biology of Schizosaccharomyces pombe. (2003)
  • Author(s): Kimura, M., Mitsuzawa, H., Ishihama.A.
  • Total Pages: 445
• [Book] Meth.Enzymol. (2003)
  • Author(s): Meates, C.F., Datwyler, S.A., Schmidt, B.D., Owens, J.Ishihama, A.
  • Total Pages: 330
  • Publisher: Academic Press, New York