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Isolation of transformation-suppressor genes by using a cDNA-expression library having sense and anti-sense inserts.

Research Project

Project/Area Number 02454145
Research Category

Grant-in-Aid for General Scientific Research (B)

Allocation TypeSingle-year Grants
Research Field General medical chemistry
Research InstitutionKobe University

Principal Investigator

KATAOKA Tohru  Kobe University, Schl. Med, Professor, 医学部, 教授 (40144472)

Co-Investigator(Kenkyū-buntansha) SUZUKI Noboru  Kobe University, Schl. Med, Researc Associate, 医学部, 助手 (00202135)
Project Period (FY) 1990 – 1991
Project Status Completed (Fiscal Year 1991)
Budget Amount *help
¥6,900,000 (Direct Cost: ¥6,900,000)
Fiscal Year 1991: ¥1,900,000 (Direct Cost: ¥1,900,000)
Fiscal Year 1990: ¥5,000,000 (Direct Cost: ¥5,000,000)
Keywordsras oncogene / RNA helicase / cDNA-expression library / reversion / リバ-ジョン / RNAhelicase / 癌抑制遺伝子
Research Abstract

1. We used artificial overexpression of cDNAs synthesized from NIH3T3 cell mRNA to induce reversion of NIH3T3 cells transformed by the activated Harvey-ras oncogene. The revertants were screened by observation under microscope for "flat" appearance. Among about 100 revertant cell lines obtained, we have chosen one revertant, Rev6-4, which showed marked decrease in its ability to form colonies in soft agar and to form tumors in nude mice, and isolated an incorporatcd cDNA from its genomic DNA by molecular cloning after PCR (polymerase chain reaction) amplification.
2. Analysis of the CDNA isolated from Rev6-4 showed that the CDNA encoded P68, which had been shown to have ATP-dependent RNA helicase activity, and that the cDNA appeared to be over-expressed in an anti-sense manner in Rev6-4 cells. Concominantly, a decrease in the amount of endogenous P68 mRNA was observed. We have confirmed that the cloned Rev6-4 CDNA could induce decrease in colony formation in soft agar, in tumor formation in nude mice and in saturation density when overexpressed in the ras-transformed NIH3T3 cells in the anti-sense manner.
3. We have isolated a full-length cDNA encoding P68. When the cDNA was over-expressed in NIH3T3 cells in a sense direction, formation of foci which appeared morphologically-transformed was observed.
However, cells in the foci could not form tumors when injected into nude mice. Presently, we do not know if the P68 gene is an oncogene whose product is acting in downstream of ras. We are currently analysing a physiological function of P68 by using antibodies against it.
4. We have isolated cDNAs from two other revertants and are currently analysing their structure and examining for their ability to induce reversion.

Report

(3 results)
  • 1991 Annual Research Report   Final Research Report Summary
  • 1990 Annual Research Report
  • Research Products

    (12 results)

All Other

All Publications (12 results)

  • [Publications] Noboru Suzuki 5: "Leucine-rich repeats and caarboxyl terminus are required for interaction of yeast adenylate cyclase with RAS proteins" Proceedings of the National Academy of Sciences,USA. 87. 8711-8715 (1990)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1991 Final Research Report Summary
  • [Publications] J.Field 5: "Cloning and Characterization of CAP,the S.cerevisiae Gene Encoding the 70kd Adenylyl Cyclase-Associated Protein" Cell. 61. 319-327 (1990)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1991 Final Research Report Summary
  • [Publications] 片岡 徹: "ras蛋白とアデニル酸シクラ-ゼ" 実験医学(増刊号). 8. 165-168 (1990)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1991 Final Research Report Summary
  • [Publications] 片岡 徹: "がん遺伝子と抑制遺伝子(第3章分担執筆)" 東京大学出版会, 143 (1991)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1991 Final Research Report Summary
  • [Publications] N. Suzuki, et. al.,: "Leucine-rich repeats and carboxyl terminus are required for interaction of yeast adenylate cyclase with RAS proteins." Proc. Natl. Acad. Sci. USA.87. 8711-8715 (1990)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1991 Final Research Report Summary
  • [Publications] J. Field, et. al.,: "Cloning and characterization of CAP, the S. cerevisiae gene encoding the 70 kd adenylate cyclase-associated protein." Cell. 61. 319-327 (1990)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1991 Final Research Report Summary
  • [Publications] T. KATAOKA: "ras protein and adenylate cyclase" Jikken Igaku. 8. 165-168 (1990)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1991 Final Research Report Summary
  • [Publications] T. KATAOKA: Tokyo Univ. Press. Oncogenes and anti-oncogenes (third chapter only), 143 (1991)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1991 Final Research Report Summary
  • [Publications] 片岡 徹: "がん遺伝子と抑制遺伝子(第3章分担執筆)" 東京大学出版会, 143 (1991)

    • Related Report
      1991 Annual Research Report
  • [Publications] Noboru Suzukiら: "Leucineーrich repeats and carboxyl terminus are required for interaction of yeast adenylate cyclase with <RAS>___ー proteins." Proceedings of the National Academy of Sciences,USA. 87. 8711-8715 (1990)

    • Related Report
      1990 Annual Research Report
  • [Publications] J.Fieldら: "Cloning and Characterization of <CAP>___ー,the S__ー.<cerevisiae>___ー Gene Encoding the 70 Kd Adenylyl CyclaseーAssociated protein" Cell. 61. 319-327 (1990)

    • Related Report
      1990 Annual Research Report
  • [Publications] 片岡 徹: "<ras>___ー蛋白とアデニル酸シクラ-ゼ" 実験医学(増刊号). 8. 165-168 (1990)

    • Related Report
      1990 Annual Research Report

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Published: 1990-04-01   Modified: 2016-04-21  

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