Juvenile hormone receptor and molecular mechanism of gene expresion

保幼激素受体及基因表达的分子机制

• 批准号: 08456171
• 负责人: CHINZEI Yasuo
• 金额: $ 4.8万
• 依托单位: Mie University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B)
• 财政年份: 1996
• 资助国家: 日本
• 起止时间: 1996 至 1997
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-08456171/
• 关键词: Juvenile hormone; cyanoprotein; gene expression; transferrin; differential display; fumin; Vitellogenin; シアノプロテイン; 貯蔵蛋白質; 転写調節; 転写調節因子; cDNAクローニング; ゲルシフトアッセイ

To study the molecular mechanism of gene expression regulation by insect juvenile hormone (JH), we have analyzed JH responsive genes by mRNA Differential Display in the bean bug, Riptortus clavatus. JH analog, methoprene, was applied on the diapause female adults and extracted RNA from the fat body. We found many cDNA bands which showed differnence between JHA and solvent acetone treated animals in mRNA Differential Display. We picked up the cDNA which were induced or repressed cDNA by JH treatment. Using these cDNA we performed Northern Blot analyzes for JH or acetone treated fat body RNA,and identified two JH inducible genes (JI-1and JI-2) and three JH repressible genes (JR-! , JR-2 and JR-3). The analysis of the partial sequeces of these cDNA and data base homology search indicated that JI-1 and JI-2 had high homology to cyanoprotein-a and vitellogenin, respectively, which we have showed hemolymph storage hexamer and egg yolk protein precursor. JR-1 was showed high homology to transferrin in vertebrate and insects. On the other hand we could not find any genes which have homology to JR-2 and JR-3 and concluded these gene are novel. We cloned and sequenced full length of cDNA of JI-2 and JR-1 and analyzed the changes of mRNA level after JH tretment by NOrthern Blot analyzes. We also cloned genomic DNA and analyzed the upstream regulatory gene structure of JI-1 (CP-a), JI-2 (Vg) and JR-1 (Transferrin). Now three JH inducible and three repressible genes are available to compare the regulation of expression by JH in the same insect under the same hormonal condition.

为了研究昆虫保幼激素（JH）对基因表达调控的分子机制，采用mRNA差异显示技术对棒条纹天牛（Riptortus clavatus）中的保幼激素应答基因进行了分析。将JH类似物甲氧普烯应用于滞育雌成虫，从脂肪体中提取RNA。在mRNA差异显示中，我们发现JHA和溶剂丙酮处理的动物之间存在许多差异的cDNA条带。提取经JH诱导或阻遏的cDNA。利用这些cDNA，我们对JH或丙酮处理的脂肪体RNA进行了北方印迹分析，并鉴定了两个JH诱导基因（JI-1和JI-2）和三个JH抑制基因（JR-1和JR-2）。、JR-2和JR-3）。对这些cDNA的部分序列分析和数据库同源性搜索表明，JI-1和JI-2分别与氰蛋白-a和卵黄蛋白原具有较高的同源性，这两个基因分别为血淋巴贮存六聚体和卵黄蛋白前体。JR-1与脊椎动物和昆虫的转铁蛋白有很高的同源性。另一方面，我们没有发现与JR-2和JR-3有同源性的基因，因此认为这些基因是新的。克隆JI-2和JR-1基因的全长cDNA并进行序列测定，用北方分析JH处理后JI-2和JR-1基因mRNA水平的变化。我们还克隆了基因组DNA并分析了JI-1（CP-α）、JI-2（Vg）和JR-1（Transferrin）的上游调控基因结构。目前已有三个JH诱导基因和三个抑制基因可用于比较JH在相同激素条件下对同一昆虫表达的调控。

项目成果

Hiroyuki Matsuoka, Hisashi Matsubara, Philomene Waidhet, Katsuhiko Ando and Yasuo Chinzei: "Depletion of salivary gland proteins in Anopheles stephenis (Diptera : Culicidae) on blood feeding, and induction of antibodies to the proteins in mice being fed."

Hiroyuki Matsuoka、Hisashi Matsubara、Philomene Waidhet、Katsuhiko Ando 和 Yasuo Chinzei：“吸血时斯氏按蚊（双翅目：蚊科）唾液腺蛋白的消耗，以及在喂食的小鼠中诱导针对该蛋白的抗体。”

Masao Yuda, Jianxin Sun, Masahiro Yamaguchi, Katsuhiko Ando and Yasuo Chinzei: "Identification and activities of anticoagulants in the salivary glands of the blood sucking kissing bug, Rhodnius prolixus." Med.Entomol.Zool.47. 263-272 (1996)

Masao Yuda、Jianxin Sun、Masahiro Yamaguchi、Katsuhiko Ando 和 Yasuo Chinzei：“吸血接吻虫（Rhodnius prolixus）唾液腺中抗凝剂的鉴定和活性。”

Jianxin Sun, et. al.: "Purification, characterization and cDNA cloning of a novel anticoagulant of the intrinsic pathway, (prolixin-S), from salivary glands of the blood sucking bug, Rhodnius prolixus." Thrombosis and Haemostasis. 75. 573-577 (1996)

孙建新,等.

P.Waidhet-Kouadio et al.: "Purification and characterization of a thrombin inhibitor from the salivary glands of a malarian vector mosquito,Anopheles stephensi." Biochemi Biophy.Acta.(in press). (1998)

P.Waidhet-Kouadio 等人：“从疟疾媒介蚊子斯蒂芬斯按蚊的唾液腺中纯化和表征凝血酶抑制剂。”

Masao Yuda, Makoto Hirai, Ken Miura, Haruo Matsumura, Katsuhiko Ando and Yasuo Chinzei: "cDNA cloning, expression and characterization of nitric-oxide synthase from the salivary glands of the blood-sucking insect, Rhodnius prolixus." Eur.J.Biochem.242. 80

Masao Yuda、Makoto Hirai、Ken Miura、Haruo Matsumura、Katsuhiko Ando 和 Yasuo Chinzei：“吸血昆虫 Rhodnius prolixus 唾液腺一氧化氮合酶的 cDNA 克隆、表达和表征。”