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ANALYSES OF FUNCTIONS OF DNA REPLICATION REGULATORY FACTORS BY THE DROSOPHILA EYE-IMAGINAL DISC SPECIFIC EXPRESSION SYSTEM

Research Project

Project/Area Number 09680640
Research Category

Grant-in-Aid for Scientific Research (C)

Allocation TypeSingle-year Grants
Section一般
Research Field Functional biochemistry
Research InstitutionAICHI CANCER CENTER

Principal Investigator

YAMAGUCHI Masamitsu  AICHI CANCER CENTER,RESEARCH INSTITUTE LABORATORY OF CELL BIOLOGY, SECTION HEAD, 生物学部, 室長 (00182460)

Co-Investigator(Kenkyū-buntansha) HIROSE Fumiko  AICHI CANCER CENTER,RESEARCH INSTITUTE LABORATORY OF CELL BIOLOGY, SENIOR RESEAR, 生物学部, 主任研究員 (60208882)
Project Period (FY) 1997 – 1998
Project Status Completed (Fiscal Year 1998)
Budget Amount *help
¥3,700,000 (Direct Cost: ¥3,700,000)
Fiscal Year 1998: ¥1,000,000 (Direct Cost: ¥1,000,000)
Fiscal Year 1997: ¥2,700,000 (Direct Cost: ¥2,700,000)
KeywordsDROSOPHILA / TRANSGENICFLY / DREF / E2F / GAL4 / TRANSCRIPTION FACTOR / DNA複製 / 複眼原基 / Pエレメント / 形態形成 / アポトーシス / Gal4
Research Abstract

Transgenic Drosophila lines expressing GAL4 specifically in the eye-imaginal discs were established. DREF cDNA and its fragments containing conserved regions 1,2 or 3 (CR1, CR2 or CR3) were ligated to the promoter containing GAL4-binding sites (UAS) in the P-element vector, and transgenic fly lines were established with these plasmid DNAs. These trnsgenic lines were utilized to examine effects of overexpression of DREF and its derivatives on eye development.
Overexpression of the wild type DREF in eye imaginal discs caused abnormal eye morphology (rough eye phenotype). The rough eye phenotype is likely caused by the ectopic induction of DNA synthesis and apoptosis in the eye-imaginal disc cells. Since it is reported that overexpression of the transcription factor E2F can induce ectopic DNA synthesis, we have crossed the DREF-overexpressing flies with the E2F mutant flies. Half-reduction of the E2F gene copy number effectively suppressed the rough eye phenotype induced by overexpression of DREF, suggesting that DREF functions upstream of the E2F gene. Furthermore, we have cloned the E2F gene and found the three DRE-related sequences in the promoter region of the E2F gene. DREF specifically binds to these DRE-related sequences of the E2F gene promoter in vitro. Detailed analyses in vitro and in vivo demonstrated that DREF can activate the E2F gene promoter.
In addition, overexpression of CRland CR3 of DREF in the eye imaginal discs also caused severe rough eye phenotype, In the eye discs of these flies, cells behind the morphogenetic furrow appeared to be arrested in G1 phase. Probably, CRland CR3 inhibited function of endogenous DREF in a dominant negative fashion to inhibit cells entering S phase.

Report

(3 results)
  • 1998 Annual Research Report   Final Research Report Summary
  • 1997 Annual Research Report
  • Research Products

    (26 results)

All Other

All Publications (26 results)

  • [Publications] Hayashi, S: "Kinase-independent activity of Cd2/CyclinA prevents sphase in the…" Genes to Cells. in press.

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1998 Final Research Report Summary
  • [Publications] Taahashi, Y.: "Identification of three conserved regions in the DREF…" Nucleic Acids Res.27. 510-516 (1999)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1998 Final Research Report Summary
  • [Publications] Sasaki, T.: "Specification of initiation regions of DNA replication in" Mol.Cell.Biol.19. 547-555 (1999)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1998 Final Research Report Summary
  • [Publications] Sawado, T.: "dE2F2, a novel E2F-family transcription factor in" Biochem.Biophys.Res.Commun.251. 409-415 (1998)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1998 Final Research Report Summary
  • [Publications] Sawado, T.: "The DNA replicaiton-related element(DRE)/DRE-binding" J.Biol.Chem.273. 26042-26051 (1998)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1998 Final Research Report Summary
  • [Publications] Kuge, M.: "Use of a fusion protein to obtain crystals suitable" Protein Science. 6. 1783-1786 (1997)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1998 Final Research Report Summary
  • [Publications] Ryu, J.-R., Choi, T.-Y., Kwon, E.-J., Lee, W.-H., Nishida, Y., Hayashi, Y., Matsukage, A., Yamaguchi, M.and Yoo, M.-A.: "Transcriptional regulation of the Drosophila raf protooncogene by the DNA replication-related element (DRE)/DRE-binding factor (DREF) system." Nucleic Acids Res.24. 794-799 (1997)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1998 Final Research Report Summary
  • [Publications] Hayashi, Y., Hirose, F., Nishimoto, Y., Shiraki, M., Yamagishi, M., Matsukage, M.and Yamaguchi, M.: "Identification of CFDD (common regulatory factor for DNA replication and DREF gene) and role of its binding site in regulation of the proliferating cell nuclear antigen gene promoter." J.Biol.Chem.272. 22848-22858 (1997)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1998 Final Research Report Summary
  • [Publications] Yamaguchi, M., Hayashi, Y., Hirose, F., Nishimoto, Y.and Matsukage, A.: "Distinct roles of E2F recognition sites as positive or negative elemens in regulation of the DNA polymerase alpha 180 kDa catalytic subunit gene promoter during Drosophila development." Nucleic Acids Res.25. 3847-3854 (1997)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1998 Final Research Report Summary
  • [Publications] Takahashi, Y., Yamaguchi, M., Hirose, F., Kobayashi, J., Miyajima S.and Matsukge, A.: "Involvement of the DNA replication-related element (DRE) and DRE-bindig factor (DREF) in transcriptional regulation of the Bombyx mori PCNA gene." J.Biochem.122. 1215-1223 (1997)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1998 Final Research Report Summary
  • [Publications] Ohno, K., Hirose, F., Inoue, Y.H., Takisawa, H., Mimura, S., Hashimoto, Y., Kiyono, T., Nishida, Y.and Matsukage, A.: "cDNA cloning and expression during development of Drosophila melanogaster MCM3, MCM6 and MCM7" Gene. 217. 177-186 (1997)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1998 Final Research Report Summary
  • [Publications] Kuge, M., Fujii, Y., Shimizu, T., Hirose, F., Matsukage, A.and Hakoshima, T.: "Use of a fusion protein to obtain crystals suitable for X-ray analysis : Crystalization of a GST-fused protein containing the DNA-binding domain of DNA replication-related element binding factor, DREF" Protein Science. 6. 1783-1786 (1997)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1998 Final Research Report Summary
  • [Publications] Sawado, T., Hirose, F., Takahashi, Y., Sasaki, T., Shinomiya, T., Sakaguchi, K., Matsukage, A.and Yamaguchi, M.: "The DNA replication-related element (DRE)/DRE-binding factor system is a transcriptiona regulator of the Drosophila E2F gene" J.Biol.Chem.273. 26042-26051 (1998)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1998 Final Research Report Summary
  • [Publications] Sawado, T., Yamaguchi, M., Nishimoto, Y., Ohno, K., Sakaguchi, K.and Matsukage, A.: "dE2F2, a novel E2F-family transcription factor in Drosophila melanogaster." Biochem.Biophys.Res.Commun.251. 409-415 (1998)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1998 Final Research Report Summary
  • [Publications] Sasake, T., Sawado, T., Yamaguchi, M.and Shinomiy, T.: "Specification of initiation regions of DNA replication in the 65-kb DNA pol alpha-dE2F locus of Drosophila melanogaster during embryogenesis and in cultured cells." Mol.Cell.Biol.19. 547-555 (1999)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1998 Final Research Report Summary
  • [Publications] Takahashi, Y., Hirose, F., Matsukage, A.and Yamaguchi, M.: "Identification of three conserved regions in the DREF transcription factors from Drosophila melanogaster and Drosophila virilis." Nucleic Acids Res.27. 510-516 (1999)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1998 Final Research Report Summary
  • [Publications] Hayashi, S.and Yamaguchi, M.: "Kinase-independent activity of Cdc2/Cyclin A prevents S phase in the Drosophila cell cycle." Genes to Cells. (in press).

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1998 Final Research Report Summary
  • [Publications] Hayashi,S.: "Kinase-independent achvity of Cdc2/CyclinA prevents Sphae" Genes to Cells. (in press).

    • Related Report
      1998 Annual Research Report
  • [Publications] Takahashi,Y.: "Identification of three conserved regions in the DREF" Nucleic Acids Res.27. 510-516 (1999)

    • Related Report
      1998 Annual Research Report
  • [Publications] Sasaki,T.: "Specification of inifiation regions of DNA replication in" Mol.Cell.Biol.19. 547-555 (1999)

    • Related Report
      1998 Annual Research Report
  • [Publications] Sawado,T.: "The DNA replication-related element(DRE)/DRE-binding" J.Biol.Chem.273. 26042-26051 (1998)

    • Related Report
      1998 Annual Research Report
  • [Publications] Sawado,T.: "dE2F2,a novel E2F-family transcription factor in" Biochem.Biophys.Res.Commun.251. 409-415 (1998)

    • Related Report
      1998 Annual Research Report
  • [Publications] Ryu,-J.-R.et al.: "Transcriptional regulation of the Drosophila raf protooncogene by ...." Nucleic Acids Res.24. 794-799 (1997)

    • Related Report
      1997 Annual Research Report
  • [Publications] Hayashi,Y.et al.: "Identification of CFDD and role of its binding site in ...." J.Biol.Chem.272. 22848-22858 (1997)

    • Related Report
      1997 Annual Research Report
  • [Publications] Yamaguchi,M.et al.: "Distinct roles of E2F recognition sites as positive or negative ..." Nucleic Acids Res.25. 3847-3854 (1997)

    • Related Report
      1997 Annual Research Report
  • [Publications] Takahashi,Y.et al.: "Involvement of the DNA replication-related element ...." J.Biochem.122. 1215-1223 (1997)

    • Related Report
      1997 Annual Research Report

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Published: 1997-04-01   Modified: 2016-04-21  

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