Nuclear function and DNA topoisomerases

核功能和 DNA 拓扑异构酶

• 批准号: 09558095
• 负责人: KIKUCHI Akihiko
• 金额: $ 3.33万
• 依托单位: Nagoya University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B).
• 财政年份: 1997
• 资助国家: 日本
• 起止时间: 1997 至 2000
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-09558095/
• 关键词: DNA topoisomerase; Monoclonal Antibody; Epitopes; Cell Division Cycle; Signal sequence; Nuclear localization; Diagnosis and therapy; Mycology; 機能領域; たんぱく分解; 細胞内局在化; 酵母; DNAポイソメラーゼ; 活性阻害; 核移行シグナル; 種特異性; クリーバブル・コンプレックス; 大腸菌による発現; トポIIα・トポI

(1) Epitope analysis of anti-DNA topoisomerase II monoclonal antibodyWe established over 100 hybridoma cells secreting anti-DNA topoisomerase II (topo II). We selected some and determined each epitope of oligopeptide level. One of them (6H8) can recognize topo II of every eucaryotes and another (8D2) is specific to proliferating cells and useful for diagnosis for neoplastic cells.(2) Analysis of topoisomerase of filamentous fungi and yeast.We cloned tope II gene of Aspergillus, Penicillium and Candida and analyzed nuclear localization signals at C-terminus region by Ala scanning method. From their sequence data we developed rapid PCR analysis for diagnosis test.(3) Novel site of localization of human topo II in nuclear regionIn addition to nuclear localization signal of human topo IIβ, we detected a novel signal sequence for the distinct localization site close to nucleolus. This signal is also found in several proteins such as RNA binding proteins and NPI46 (nucleolar peptidyl prolyl cis trans isomerase).(4) The monoclonal antibodies against human DNA topoisomerase IIαextensively used and characterized throughout this research project was filed for patent application by Mitsubishi-Kagaku and now comercialized for the diagnostic reagent of malignant tumor cells.

(1)抗DNA拓扑异构酶II单克隆抗体的表位分析我们建立了100多个分泌抗DNA拓扑异构酶II(topo II)的杂交瘤细胞，并测定了其中一个(6H8)可以识别寡肽水平的每个表位。每种真核生物和另一种真核生物 (8D2) 对增殖细胞具有特异性，可用于肿瘤细胞的诊断。(2) 拓扑异构酶分析我们克隆了曲霉属、青霉属和念珠菌属的tope II基因，并通过Ala扫描方法分析了C末端区域的核定位信号，根据它们的序列数据，我们开发了用于诊断测试的快速PCR分析。人拓扑II在核区的定位除了人拓扑IIβ的核定位信号外，我们还检测到靠近核仁的独特定位位点的新信号序列。 RNA 结合蛋白和 NPI46（核仁肽基脯氨酰顺反异构酶）等多种蛋白质。(4) 在整个研究项目中广泛使用和表征的抗人 DNA 拓扑异构酶 IIα 的单克隆抗体已由三菱化学申请专利，现已商业化恶性肿瘤细胞诊断试剂。

项目成果

K.MATSUMOTO et al.: "Sperm chromatin decondensation by template activating factor I"Mol.Cell.Biol.. 19. 6940-6952 (1999)

K.MATSUMOTO 等人：“通过模板激活因子 I 进行精子染色质去浓缩”Mol.Cell.Biol.. 19. 6940-6952 (1999)

Y.OKADA,Y.ITO,A.KIKUCHI,Y.NIMURA,S.YOSHIDA and M.SUZUKI: "Assignment of functional aminoacids around the active site of human DNA topoisomerase IIα"J.Biol.Chem.. 275,. 24630-24638 (2000)

Y.OKADA、Y.ITO、A.KIKUCHI、Y.NIMURA、S.YOSHIDA 和 M.SUZUKI：“人类 DNA 拓扑异构酶 IIα 活性位点周围的功能氨基酸的分配”J.Biol.Chem.. 275，。 -24638 (2000)

K.SUGIMOTO et al.: "Temporal and spatial distribution of DNA topoisomerase II alters during proliferation,differenciation and apoptosis in HL-60 cells" Blood. 91. 1407-1417 (1998)

K.SUGIMOTO 等人：“HL-60 细胞增殖、分化和凋亡过程中 DNA 拓扑异构酶 II 的时间和空间分布发生变化”血液。

N.ADACHI, M.MIYAIKE, S.KATO, R.KANAMARU, H.KOYAMA and A.KIKUCHI: "Cellular distribution of mammalian DNA topoisomerase II is determined by its catalytically dispensable C-terminal domain"Nucl.Acids Res. 25. 3135-3142 (1997)

N.ADACHI、M.MIYAIKE、S.KATO、R.KANAMARU、H.KOYAMA 和 A.KIKUCHI：“哺乳动物 DNA 拓扑异构酶 II 的细胞分布是由其催化可分配的 C 末端结构域决定的”Nucl.Acids Res。

Y.Shimizu,T.Akashi,A.Okuda,A.Kikuchi,K.Fukui: "NBP1 (Nap1 binding protein1), an essential gene for G2/M transition of S.cerevisiae, encodes a protein of distinct sub-nuclear localization"GENE. 246,. 395-404 (2000)

Y.Shimizu、T.Akashi、A.Okuda、A.Kikuchi、K.Fukui：“NBP1（Nap1 结合蛋白 1）是酿酒酵母 G2/M 转变的必需基因，编码一种具有不同亚核定位的蛋白质