Identification and functional analyses of genetic interactants with DREF, a master regulator of DNA replication genes

2005 Fiscal Year Final Research Report Summary

• Project/Area Number: 16370004
• Research Category: Grant-in-Aid for Scientific Research (B)
• Allocation Type: Single-year Grants
• Section: 一般
• Research Field: Genetics/Genome dynamics
• Research Institution: Kyoto Institute of Technology
• Principal Investigator: YAMAGUCHI Masamitsu Kyoto Institute of Technology, Faculty of Textile Science, Professor, 繊維学部, 教授 (00182460)
• Project Period (FY): 2004 – 2005
• Keywords: transcription factor / transcriptional regulation / translation initiation factor / ubiquitin ligase / Drosophila melanogaster / genetics / chromatin immunoprecipitation / RNAi

Research Abstract

The DNA replication-related element-binding factor (DREF) has been suggested to be involved in regulation of DNA replication and proliferation-related genes in Drosophila. Over-expression of DREF induced ectopic DNA synthesis and apoptosis, and inhibited the photoreceptor cell differentiation in eye imaginal discs and adult flies exhibited a severe rough eye phenotype. We used DREF over-expressing fly lines as a tool to screen mutations that modify the rough eye phenotype. We identified 20 lines including eIF4a and skpA gene mutations that suppressed and 6 lines that enhanced the rough eye phenotype when they were heterozygous for the mutations.
The eIF4a gene carries three DRE sequences in its 5'-flanking region. Luciferase transient expression assays in cultured Drosophila cells with eIF4a promoter-luciferase fusion plasmids and chromatin immunoprecipitation assays using anti-DREF antibody revealed that DREF directly regulates expression of the eIF4a gene. Analyses with transgenic fli es carrying eIF4a promoter-lacZ fusion genes also support this conclusion. Since it is reported that eIF4a protein is associated with growth by regulating protein translation, regulation of eIF4a gene expression by DREF would be important for regulation of cell proliferation.
SKPa is component of a Drosophila SCF complex that functions in combination with the ubiquitin conjugating enzyme UbcD1. DRE sequences were found in skpA promoter. Both luciferase transient expression assays in cultured Drosophila S2 cells using skpA promoter-luciferase fusion plasmids and anti-lacZ immunostaining of various tissues from transgenic third instar larvae carrying the skpA promoter-lacZ fusion genes provided supportive evidence. Furthermore, anti-SKPa immunostaining of eye imaginal discs from flies over-expressing DREF showed ectopic expression of protein in the region posterior to the morphogenetic furrow where DREF is over-expressed. Knockdown of DREF in some tissues almost completely abrogated the skpA gene expression. These findings indicate that the Drosophila skpA gene is a novel target of the transcription factor DREF.

Research Products

• [Journal Article] Drosophila Damaged DNA Binding Protein 1 contributes to genome stability in somatic cells. (2006)
  • Author(s): Shimanouchi, K., Takata, K., Yamaguchi, M., Murakami, S., Ishikawa, G., Takeuchi, R., Kanai, Y., Ruike, T., Nakamura, R., Abe, Y., Sakaguchi, K
  • Journal Title: J.Biochem. 139 Pages: 51-58
  • Description: 「研究成果報告書概要(和文)」より
• [Journal Article] Antagonistic regulation of the Drosophila PCNA gene promoter by DREF and Cut. (2006)
  • Author(s): Seto, H., Hayashi, Y., Kwon, E., Taguchi, O., Yamaguchi, M
  • Journal Title: Genes to Cells 11 Pages: 499-512
  • Description: 「研究成果報告書概要(和文)」より
• [Journal Article] Drosophila Damaged DNA Binding Protein 1 contributes to genome stability in somatic cells. (2006)
  • Author(s): Shimanouchi, K., Takata, K., Yamaguchi, M., Murakami, S., Ishikawa, G., Takeuchi, R., Kanai, Y., Ruike, T., Nakamura, R., Abe, Y., Sakaguchi, K.
  • Journal Title: J.Biochem. 139 Pages: 51-58
  • Description: 「研究成果報告書概要(欧文)」より
• [Journal Article] Antagonistic regulation of the Drosophila PCNA gene promoter by DREF and Cut. (2006)
  • Author(s): Seto, H., Hayashi, Y., Kwon, E., Taguchi, O., Yamaguchi, M.
  • Journal Title: Genes to Cells 11 Pages: 499-512
  • Description: 「研究成果報告書概要(欧文)」より
• [Journal Article] Alternative splicing regulates the transcriptional activity of Drosophila heat shock transcription factor in response to heat/cold stress. (2005)
  • Author(s): Fujikake, N., Nagai, Y., Popiel H.A., Kano, H., Yamaguchi, M., Toda, T.
  • Journal Title: FEBS letter 579 Pages: 3842-3848
  • Description: 「研究成果報告書概要(和文)」より
• [Journal Article] Transcriptional regulation of the Drosophila orc2 gene by the DREF pathway. (2005)
  • Author(s): Okudaira, K., Ohno, K., Yoshida, H., Asano, M., Hirose, F., Yamaguchi, M
  • Journal Title: Biochem.Biophys.Acta. 1732 Pages: 23-30
  • Description: 「研究成果報告書概要(和文)」より
• [Journal Article] Transcriptional regulation of the Drosophila orc2 gene by the DREF pathway. (2005)
  • Author(s): Okudaira, K., Ohno, K., Yoshida, H., Asano, M., Hirose, F., Yamaguchi, M.
  • Journal Title: Biochem.Biophys.Acta. 1732 Pages: 23-30
  • Description: 「研究成果報告書概要(欧文)」より
• [Journal Article] Redox regulation of DNA binding activity of DREF in Drosophila. (2004)
  • Author(s): Choi, T.-Y., Park, S.Y., Kang, H.-S., Cheong, J.-H., Kim, H.-D., Lee, B.L., Hirose, F., Yamaguchi, M., Yoo, M.-A.
  • Journal Title: Biochem.J. 378 Pages: 833-838
  • Description: 「研究成果報告書概要(和文)」より
• [Journal Article] Drosophila damaged DNA binding protein 1 (D-DDB1) is an essential factor for development. (2004)
  • Author(s): Takata, K., Yoshida, H., Yamaguchi, M., Sakaguchi, K.
  • Journal Title: Genetics 168 Pages: 855-865
  • Description: 「研究成果報告書概要(和文)」より
• [Journal Article] Genetic Link between p53 and genes required for formation of the zonula adeherens junction. (2004)
  • Author(s): Yamaguchi, M., Hirose, F., Inoue Y., H., Ohno, K., Yoshida, H., Hayashi, Y., Deak, P., Matsukage, A
  • Journal Title: Canser Sci. 95 Pages: 436-441
  • Description: 「研究成果報告書概要(和文)」より
• [Journal Article] Phenotypic stability of the P-M system in wild populations of Drosophila melanogaster. (2004)
  • Author(s): Itoh, M., Fukui, T., Kitamura, M., Uenoyama, T., Watada, M., Yamaguchi, M
  • Journal Title: Genes Genet.Syst. 79 Pages: 9-18
  • Description: 「研究成果報告書概要(和文)」より
• [Journal Article] Transcriptional regulation of the Drosophila caudal homeobox gene by the DRE/DREF system. (2004)
  • Author(s): Choi, Y.-J., Choi, T.-Y., Yamaguchi, M., Matsukage, A., Kim.Y.-S., Yoo, M.-A
  • Journal Title: Nucleic Acids Res. 32 Pages: 3734-3742
  • Description: 「研究成果報告書概要(和文)」より
• [Journal Article] DREF is required for EGFR signaling during Drosophila wing vein development. (2004)
  • Author(s): Yoshida, H., Kwon, E., Hirose, F., Otsuki, K., Yamada, M., Yamaguchi, M
  • Journal Title: Genes to Cells 9 Pages: 935-944
  • Description: 「研究成果報告書概要(和文)」より
• [Journal Article] Armadillo/Pangolin regulates PCNA and DREF promoter activities. (2004)
  • Author(s): Kwon E., Hayashi, Y., Otuski, K., Hirose, F., Nishida, Y., Yoo, M.-A., Yamaguchi, M.
  • Journal Title: Biochem.Biophys.Acta. 1676 Pages: 256-262
  • Description: 「研究成果報告書概要(和文)」より
• [Journal Article] Damaged DNA binding protein 1 in Drosophila defense reactions. (2004)
  • Author(s): Takata, K., Shimanouchi, K., Yamaguchi, M., Murakami, S., Ishikawa, G., Takeuchi, R., Kanai, Y., Ruike, T., Nakamura, R., Abe, Y., Sakaguchi, K
  • Journal Title: Biochem.Biophys.Res.Commun. 323 Pages: 1024-1031
  • Description: 「研究成果報告書概要(和文)」より
• [Journal Article] Characterization of dRFX2, a novel RFX family protein in Drosophila. (2004)
  • Author(s): Otsuki, K., Hayashi, Y., Kato, M., Yoshida, H., Yamaguchi, M
  • Journal Title: Nucleic Acids Res. 32 Pages: 5636-5648
  • Description: 「研究成果報告書概要(和文)」より
• [Journal Article] Genetic Link between p53 and genes required for formation of the zonula adherens junction. (2004)
  • Author(s): Yamaguchi, M., Hirose, F., Inoue Y.H., Ohno, K., Yoshida, H., Hayashi, Y., Deak, P., Matsukage, A.
  • Journal Title: Cancer Sci. 95 Pages: 436-441
  • Description: 「研究成果報告書概要(欧文)」より
• [Journal Article] Phenotypic stability of the P-M system in wild populations of Drosophila melanogaster. (2004)
  • Author(s): Itoh, M., Fukui, T., Kitamura, M., Uenoyama, T., Watada, M., Yamaguchi, M.
  • Journal Title: Genes Genet.Syst. 79 Pages: 9-18
  • Description: 「研究成果報告書概要(欧文)」より
• [Journal Article] Transcriptional regulation of the Drosophila caudal homeobox gene by the DRE/DREF system. (2004)
  • Author(s): Choi, Y.-J., Choi, T.-Y., Yamaguchi, M., Matsukage, A., Kim.Y.-S., Yoo, M.-A.
  • Journal Title: Nucleic Acids Res. 32 Pages: 3734-3742
  • Description: 「研究成果報告書概要(欧文)」より
• [Journal Article] DREF is required for EGFR signaling during Drosophila wing vein development. (2004)
  • Author(s): Yoshida, H., Kwon, E., Hirose, F., Otsuki, K., Yamada, M., Yamaguchi, M.
  • Journal Title: Genes to Cells 9 Pages: 935-944
  • Description: 「研究成果報告書概要(欧文)」より
• [Journal Article] Armadillo/Pangolin regulates PCNA and DREF promoter activities. (2004)
  • Author(s): Kwon E., Hayashi, Y., Otsuki, K., Hirose, F., Nishida, Y., Yoo, M.-A., Yamaguchi, M.
  • Journal Title: Biochem.Biophys.Acta. 1679 Pages: 256-262
  • Description: 「研究成果報告書概要(欧文)」より
• [Journal Article] Damaged DNA binding protein 1 in Drosophila defense reactions. (2004)
  • Author(s): Takata, K., Shimanouchi, K., Yamaguchi, M., Murakami, S., Ishikawa, G., Takeuchi, R., Kanai, Y., Ruike, T., Nakamura, R., Abe, Y., Sakaguchi, K.
  • Journal Title: Biochem.Biophys.Res.Commun. 323 Pages: 1024-1031
  • Description: 「研究成果報告書概要(欧文)」より
• [Journal Article] Characterization of dRFX2, a novel RFX family protein in Drosophila. (2004)
  • Author(s): Otsuki, K., Hayashi, Y., Kato, M., Yoshida, H., Yamaguchi, M.
  • Journal Title: Nucleic Acids Res. 32 Pages: 5636-5648
  • Description: 「研究成果報告書概要(欧文)」より
• [Book] 増殖関連遺伝子の制御機構.発生における細胞増殖制御(竹内隆, 岸本健雄編 (2004)
  • Author(s): 山口政光, 加藤容子
  • Total Pages: 9
  • Publisher: シュプリンガーフェアラーク東京
  • Description: 「研究成果報告書概要(和文)」より
• [Book] ショウジョウバエのヘテロクロマチン形成.エピジェネテイクス(佐々木裕之編) (2004)
  • Author(s): 山口政光, 西田美樹
  • Total Pages: 10
  • Publisher: シュプリンガーフェアラーク東京
  • Description: 「研究成果報告書概要(和文)」より