多田 周右 東京大学, 薬学部, 助手 (00216970)
関 政幸 東北大学, 薬学部, 助手 (70202140)
ISHーHOROWICZ 英国癌研究基金リンカーンズ, イン・フィールド研究所, 主席研究官
ISH-HOROWICZ David Imperial Cancer Research Fund Lincoln's Inn Fields
SEKI Masayuki Faculty of Pharmaceutical Sciences, Tohoku University
|Budget Amount *help
¥7,100,000 (Direct Cost : ¥7,100,000)
Fiscal Year 1996 : ¥3,400,000 (Direct Cost : ¥3,400,000)
Fiscal Year 1995 : ¥3,700,000 (Direct Cost : ¥3,700,000)
DNA helicase Q1 is a human homologue of the Escherichia coli DNA helicase RecQ,which we found in human cells. The existence of a RecQ homologue in human cells indicated that eukaryotic cells had a DNA repair-recombination system involving eukaryotic RecQ.In this study, we intended to clarify functions of eukaryoric RecQ in the novel DNA repair-recombination system and in the process of development using yeast, mouse, and fly.
1 Analysis of the function of yeast RecQ (Sgs1) with SGS1 gene disruptants
We cloned a yeast RecQ gene by using human DNA helicase Q1 cDNA as a probe and found that the cloned gene was identical to SGS1 which was cloned as a suppresser of the slow growth phenotype of topoisomerase III mutants of Saccharomyces cerevisiae. We analyzed the sensitivity of SGS1 gene disruptants to various genotoxic agents. Disruptants showed higher sensitivity to alkylating agents such as methy methanesulfonate and ethyl methanesulfonate (MMS) and hydroxyurea (HU) as compared with wild-t
ype cells. When homozygous SGS1 disruptants were transferred to sporulation medium, poor sporulation and a reduced frequency of meiotic recombination were observed. In addition, the induction of expression of SGS1 mRNA was observed when wild-type cells were treated with MMS or transferred to sporulation medium to induce meiosis. These results indicates that Sgs1 is involved in the repair of some type of DNA lesions and meiotic DNA recombination.
2 cDNA cloning of mouse DNA helicase Q1 and analysis of expression of Q1 mRNA
We cloned a cDNA encoding mouse DNA helicase Q1 and examined expression of Q1 message in various mouse tissues. The mRNA was expressed in the highest level in testis, a higher level in thymus and was poorly expressed in brain, heart, kidney, liver, lung, muscle, ovary, and spleen. When the expression of Q1 mRNA in testis was monitored after birth, an increase in the level of the mRNA was observed 14th day after birth when cells in zygotene and pachytene increased, in which DNA recombination took place.
3 cDNA cloning of Dorosophila DNA helicase Q1
We have succeeded to isolate a Dorosophila DNA helicase Q1 cDNA.However, the analysis of roles of Q1 during development by making transgenic flies has remained for future study.