Study on initiation of DNA repication in cell-free system of Xenopus eggs

• Project/Area Number: 07680762
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Single-year Grants
• Section: 一般
• Research Field: Cell biology
• Research Institution: Kumamoto University
• Principal Investigator: SHIODA Masaki Kumamoto University Fac.Science, Dept.Biological Science, Professor, 理学部, 教授 (80134526)
• Project Period (FY): 1995 – 1997
• Project Status: Completed (Fiscal Year 1997)
• Budget Amount: ¥2,300,000 (Direct Cost: ¥2,300,000); Fiscal Year 1997: ¥700,000 (Direct Cost: ¥700,000); Fiscal Year 1996: ¥800,000 (Direct Cost: ¥800,000); Fiscal Year 1995: ¥800,000 (Direct Cost: ¥800,000)
• Keywords: Xenopus laevis / DNA rreplication / cell-free DNA replication / initiator protein / license factor / initation of DNA replication / MCM / DNA untwisting protein / 細胞周期 / 胚発生 / P1蛋白質 / リン酸化 / 複製開始蛋白質 / DNAヘリカーゼ

Research Abstract

We purified a novel protein (designated to DUF) which has an activity to untwist DNA and open double strand DNA to single strand DNA from cell-free DNA synthetic system of Xenopus eggs. DUF was composed of 87 and 140 kDa polypeptides. Immunobloting exper iments using antibody against the 87 kDa polypeptide showed that behavior of DUF during a cell cycle or embryogenesis co-relates with DNA replication. Immunodepletion of DUF from the DNA synthetic system lead to inhibition of intiation of DNA replication. These results suggest that DUF participates in intiation on of DNA replication. We also surveeid protein factors which regulate intiation of DNA replication using antibody agaist proteins concerning to DNA replication (human P1Cdc46 and human P1). As results, we detected a protein complex composed of 5 prteeins containing MCM famliy which regulates intiaton of DNA replication, correspondiing too the license factor which allows DNA replication only once in a cell cycle. Further studies on the relationship between DUF and license factor are required to elucidate regulation of initiation of DNA replication.

Research Products

• [Publications] Someya, A.: "The association and involvement of the P1 protein family in a cell free extract of Xenopus eggs" Biochem.Biophys.Res.Commun.209. 823-831 (1995)
  • Description: 「研究成果報告書概要(和文)」より
• [Publications] Someya.A.: "The possible involvement of replication-related proteins with a DEAD-box-like motif in cell-free DNA replication of Xenopus eggs" Biochem.Biophys.Res.Commun.212. 1098-1106 (1995)
  • Description: 「研究成果報告書概要(和文)」より
• [Publications] Someya, A.: "The possible involvement of DNA-dependent protein kinase in the phosphorylation of P1 protein in cell-free DNA replication of Xenopus eggs" Biochem.Biophys.Res.Commun.213. 803-814 (1995)
  • Description: 「研究成果報告書概要(和文)」より
• [Publications] Murakami-Murofushi, K.: "Selective inhibition of DNA polymerase a family with chemically synthesized derivatives of PHYLPA,a unique Physarum lysophosphatidic acid" Biochim.Biophys.Acta. 1258. 57-60 (1995)
  • Description: 「研究成果報告書概要(和文)」より
• [Publications] Takayanagi, S.: "Sulfolobus hakonensis sp.nov.,anovel species of acidothermophilic Archaeon" Int.J.System.Bacteriol.46・2. 377-382 (1996)
  • Description: 「研究成果報告書概要(和文)」より
• [Publications] Yokoyama, Y.: "Enhancement of apoptosis in developing chick neural retina cells by basic fibroblast growth factor" Jounal of Neurochemistry. 68. 2212-2215 (1997)
  • Description: 「研究成果報告書概要(和文)」より
• [Publications] Yokoyama, Y.: "CPP32 activation during dolichyl phosphate-induced apoptosis in U937 leukemia cells" FEBS Letters. 412. 153-156 (1997)
  • Description: 「研究成果報告書概要(和文)」より
• [Publications] Someya, A., Shioda, M., Okuyama, A.: "The association and involvement of some member of the P1 protein family in a cell-free DNA replication of Xenopus eggs." Biochem Biophys.Res.Commun. 209. 823-831 (1995)
  • Description: 「研究成果報告書概要(欧文)」より
• [Publications] Someya, A., and Shioda, M.: "The possible involvement of replication-related proteins with a DEAD-box-like motifin in cell-free DNA replication of Xenopus eggs." Biochem Biophys.Res.Commun. 212. 1098-1106 (1995)
  • Description: 「研究成果報告書概要(欧文)」より
• [Publications] Someya, A., Take, Y., Shioda, M.: "The possible involvement of DNA-dependent protein kinase in the phosphorylation of P1 protein in cell-free DNA replication of Xenopus eggs." Biochem Biophys.Res.Commun. 213. 803-814 (1995)
  • Description: 「研究成果報告書概要(欧文)」より
• [Publications] Murakami-Murofushi, K., Kobayashi, S., Onimura, K., Matsumoto, M., Yoshida, S., Shoji, M., Murofushi, H.: "Selective inhibition of DNA polymerase-alpha family with chemically synthesized derivatives of PHYLPA,a unique Physarum lysophosphatidic acid." Biochim.Biophys.Acta. 1258. 57-60 (1995)
  • Description: 「研究成果報告書概要(欧文)」より
• [Publications] Takayanagi, S., Kawasaki, H., Sugimori, K., Yamada, T., Sugai, A., Ito, T., Yamasato, K., and Shioda, M.: "Sulfolobus hakonensis sp.nov., a novel species of acidothermophilic Archaeon." Int.J.System.Bacteriol. 46. 377-382 (1996)
  • Description: 「研究成果報告書概要(欧文)」より
• [Publications] Yokoyama, Y., Okubo, T., Ozawa, S., Nagai, F., Ushiyama, K., Kano, I., Shioda, M., Kubo, H., Takemura, M., Namiki, H., Yasugi, E., Oshima, M., Seyama, Y., and Kano, K.: "CPP32 activation during dolichyl phosphate-induced apoptosis in U937 leukemia cells." FEBS Lett.412. 153-156 (1997)
  • Description: 「研究成果報告書概要(欧文)」より
• [Publications] Yokoyama, Y., Ozawa, S., Seyama, Y., Namiki, H., Hatashi, Y., Kaji, K., Shirama, K., Shioda, M., and Kano, K.: "Enhancement of apoptosis in developing chick neural retina cells by basic fibroblast growth factor." J.Neurochem. 68. 2212-2215 (1997)
  • Description: 「研究成果報告書概要(欧文)」より
• [Publications] Yokoyama,Y.: "Enhancement of apoptosis in developing chick neural retina cells by basic fibroblast growth factor" Journal of Neurochemistry. 68. 2212-2215 (1997)
• [Publications] Yokoyama,Y.: "CPP32 activation during dolichyl phosphate-induced apoptosis in U937 leukemia cells" FEBS Letters. 412. 153-156 (1997)
• [Publications] Takayanagi,S.: "Sulfolobus hakonensis sp.nov.,a novel species of acidothermophilic Archaeon" Int.J.System.Bacteriol.46・2. 377-382 (1996)
• [Publications] Yokoyama,Y.: "Enhancement of apoptosis in developing chick neural retina cells by basic fibroblast growth factor" Journal of Neurochemistry. (印刷中). (1997)
• [Publications] Someya,A.: "The association and involvement of the P1 protein family in a cell free extract of Xenopus eggs" Biochem.Biophys.Res.Commun.209. 823-831 (1995)
• [Publications] Someya,A.: "The possible involvement of replication-related proteins with a DEAD-box-like motif in cell-free DNA replication of Xenopus eggs" Biochem.Biophys.Res.Commun.212. 1098-1106 (1995)
• [Publications] Someya,A.: "The possible involvement of DNA-dependent protein kinase in the phosphorylation of P1 protein in cell-free DNA replication of Xenopus eggs" Biochem.Biophys.Res.Commun.213. 803-814 (1995)
• [Publications] Murakami-Murofushi,K.: "Selective inhibition of DNA polymerase a family with chemically synthesized derivatives of PHYLPA,a unique Physarum lysophosphatidic acid" Biochim.Biophys.Acta. 1258. 57-60 (1995)