CHARACTERIZATION OF SENESCENCE GENES IN HUMAN CELLS

人类细胞衰老基因的特征

• 批准号: 06454675
• 负责人: AYUSAWA Dai
• 金额: $ 4.03万
• 依托单位: KIHARA INSTITUTE FOR BIOLOGICAL RESEARCH (1995-1996)The University of Tokyo (1994)
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B)
• 财政年份: 1994
• 资助国家: 日本
• 起止时间: 1994 至 1996
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-06454675/
• 关键词: SENESCENCE; IMMORTALIZATION; GENE; CHROMOSOME 7; GENE MAPPING; RADIATION HYBRID; 細部老化; 細部不死化; 細胞老化遺伝子; 情報伝達; 不死化細胞; D群不死化細胞株; cGMP-依存プロテインキナーゼ; シグナル伝達系

We have undertaken the following two projects in a given time, and obtained several new findings that contribute understanding of cellular senescence and imnrotalization in human cells.1. Identification of a senescence / immortalizing gene on human chromosome 7We made a panel of G418-resistant radiation hybrid cells form mouse A7 cells containing human chromosome 7 tagged with pSV2neo. By analysis of FISH,chromosome transfer, and Alu PCR,we selected radiation hybrids that induce senescence in immortal cell lines assigned to genetic colmnplemenation D and have a very limited amount of human DNA.The human DNA was mapped to chromosome 7 to molecularly clone the senescence gene in a YAC vector.2. Characterization of a signal transduction pathway in cellular senescenceInhibitors of cGMP-dependent protein kinases was found to block senescence induced by heat inactivation of SV40T antigen in SV40T-transformed immortal human fibroblasts. Using one of such inhibitors as a ligand for affinity chromatography followed by Western blot analysis, we have found that a protein phosphatase was induced and dephosphorylation of specific proteins occurred. The mechanism of induction of the phosphatase and the proteins dephosphorylated were examined in vino and in vitro using specific inhibitors of phosphatases and antibodies.

我们在给定的时间内进行了以下两个项目，并获得了一些新的发现，有助于理解人类细胞的衰老和永生化。人7号染色体衰老/永生化基因的鉴定我们用pSV 2neo标记人7号染色体，从小鼠A7细胞中制备了一组抗G418辐射的杂交细胞。通过荧光原位杂交、染色体转移和Alu PCR分析，筛选出了能诱导衰老的永生细胞系，并将其定位于第7号染色体，将衰老基因克隆到YAC载体中.细胞衰老信号转导途径的表征发现cGMP依赖性蛋白激酶抑制剂可阻断SV 40 T转化的永生人成纤维细胞中SV 40 T抗原热灭活诱导的衰老。使用一种这样的抑制剂作为亲和层析的配体，然后进行Western印迹分析，我们发现，蛋白磷酸酶被诱导，并发生特定蛋白的去磷酸化。磷酸酶和去磷酸化的蛋白质的诱导的机制进行了检查，在vino和体外使用磷酸酶和抗体的特异性抑制剂。

项目成果

T.Ogata et al.: "Genetic complementation of the immortal phenotype ingroup D cell lines byintroduction of chromosome 7" Jpn.J.Cancer Res.86. 35-40 (1995)

T.Ogata 等人：“通过引入 7 号染色体对 D 组细胞系中永生表型进行遗传互补”Jpn.J.Cancer Res.86。

T.Nakamura, R.Sanokawa, Y.Sasaki, D.Ayusawa, M.Oishi, and N.Mori.: "N-Shc : a neural specific adaptor molecule that mediates signaling from neutrophin / TrK to Ras / MAPK pathway." Oncogene. 13. 1111-1121 (1996)

T.Nakamura、R.Sanokawa、Y.Sasaki、D.Ayusawa、M.Oishi 和 N.Mori.：“N-Shc：一种神经特异性接头分子，介导从中性粒细胞/TrK 到 Ras/MAPK 途径的信号传导。”

M.Fujii et al.: "Inhibitors of cGMP-dependent protein kinase block senescence induced by inactivation of T antigen in SV40-transformed immortal human fibroblasts" Oncogene. 11. 627-634 (1995)

M.Fujii 等人：“cGMP 依赖性蛋白激酶抑制剂可阻止 SV40 转化的永生人类成纤维细胞中 T 抗原失活诱导的衰老”Oncogene。

鮎沢大: "ヒト細胞の不死化に関する遺伝子" 組織培養. 22. 24-2 (1996)

Dai Ayuzawa：“与人类细胞永生化相关的基因”组织培养22. 24-2 (1996)。

M.Fujii et al.: "Expression of the human c GMP-dependent protein kinase type II gene is lost upon introduction of SV40 T antigen or immortalization in human fibroblasts" FEBS letters. 375. 263-267 (1995)

M.Fujii 等人：“在人成纤维细胞中引入 SV40 T 抗原或永生化后，人 c GMP 依赖性蛋白激酶 II 型基因的表达就会丢失”FEBS 字母。