Mechanisms of recognition of non-self and initiation of defense reactions in insects

昆虫非我识别和防御反应启动机制

• 批准号: 02454019
• 负责人: ASHIDA Masaaki
• 金额: $ 4.03万
• 依托单位: Hokkaido University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for General Scientific Research (B)
• 财政年份: 1990
• 资助国家: 日本
• 起止时间: 1990 至 1992
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-02454019/
• 关键词: INSECT; HEMOLYMPH; PHENOLOXIDASE; CUTICLE; CECROPIN; DEFENSE; PROTEASE; PEPTIDOGLYCAN; 異物認識; フェノ-ル酸化酵素; ペプチドグリカン; カビ; バクテリア; フェノ-ル酸化酵素前駆体; 蚊; ゴキブリ; バッタ

The ultimate end of this research project is to study the mechanism by which foreign materials such as bacteria, fungi and parasites invading insects are recognized as non-self and how the recognition mechanism of non-self is linked to the initiation of defense reactions of insects. To reach the end the present research was carried out using two insect tissues, hemolymph and integument of the silkworm, Bombyx mori.The prophenoloxidase cascade (ProPO cascade) is present in hemolymph. The cascade is triggered by minute amounts of beta-1,3-glucan (beta G) or peptidoglycan (PG). We have shown previously that PG and betaG trigger the cascade when they bind specifically to peptidoglycan recognition protein (PGRP) and beta-1,3-glucan recognition protein (beta GRP), respectively. Although it is speculated that the recognition proteins may play a central role in the recognition of bacteria or fungi as non-self in insect hemocoel, our knowledge of the chemical structures and the mode of action o … More f the recognition proteins is very limited. We investigated the localization of the recognition proteins in hemocoel by immunochemical and immunocytochemical methods and put forward a working hypothesis on the role of the two recognition proteins in the process of recognition of bacteria or fungi as non-self in hemocoel.The components of proPO cascade include more than two serine enzyme zymogens. Experimental results so har obtained suggest that active forms of the zymogens or fragments derived from the zymogens during their activation modulate phagocytotic activity and hemokinetic activity of hemocytes. To prove this possibility unambiguously we are currently trying to purify all proPO cascade components. We recently have succeeded in purifying proBAEEase and its active form to homogeneity. It is expected that the active enzyme will be studied whether it has the effect on the phagocytotic or hemokinetic activity of hemocytes.Cuticle of insect integument is often considered to be a protective armor and chemically inert (dead) part of the body. Using axenically raised larvae of silkworm, Bombyx mori we could demonstrate that the cuticle transmit a signal to epidermal cells of integument to direct them to synthesize antibacterial peptide (cecropin) when the cuticle has bacterial cell wall component (lipopolysaccharide or peptidoglycan) on its abraded part. The antibacterial peptide is secreted to cuticle. The presence of bacterial cell wall component on abraded cuticle was shown to cause the synthesis of cecropin mRNA in fat body, but in this case no cecropin activity was detected in fat body and hemolymph. Seemingly this mRNA synthesis without immediate translation of the message indicate that cuticular abrasion with bacterial cell wall component make the the entire insect body alert to bacterial infection. Currently we are investigating the nature of signal sent to epidermal cells and fat body from the abraded part of cuticle with bacterial cell wall component. Less

该研究项目的最终目的是研究入侵昆虫的细菌、真菌和寄生虫等外来物质被识别为非自体的机制，以及非自体的识别机制如何与昆虫防御反应的启动联系起来。为了达到这个目的，本研究使用两种昆虫组织，即家蚕的血淋巴和体皮进行。原酚氧化酶级联（ProPO级联）存在于血淋巴中。该级联反应是由微量的 β-1,3-葡聚糖 (β G) 或肽聚糖 (PG) 触发的。我们之前已经证明，当 PG 和 betaG 与肽聚糖识别蛋白 (PGRP) 和 β- 特异性结合时，会触发级联反应。 1,3-葡聚糖识别蛋白（β GRP），尽管据推测，识别蛋白可能在昆虫将细菌或真菌识别为非自身的过程中发挥着核心作用。血腔，我们对识别蛋白的化学结构和作用方式的了解非常有限，我们通过免疫化学和免疫细胞化学方法研究了识别蛋白在血腔中的定位，并提出了关于识别蛋白的作用的工作假设。 proPO级联的组成部分包括两种以上的丝氨酸酶酶原，所获得的实验结果表明，proPO级联的成分是两种以上的丝氨酸酶原。酶原或源自酶原的片段在其激活过程中调节血细胞的吞噬活性和血液动力学活性，为了明确地证明这种可能性，我们目前正在尝试纯化所有 proPO 级联成分，我们最近成功地纯化了 proBAEEase 及其活性形式。预计将研究活性酶是否对血细胞的吞噬或血液动力学活性有影响。昆虫体皮通常被认为是身体的保护性盔甲和化学惰性（死亡）部分，利用蚕（Bombyx mori）的无菌饲养幼虫，我们可以证明角质层向体皮表皮细胞传递信号，指导它们合成抗菌剂。当角质层的磨损部分有细菌细胞壁成分（脂多糖或肽聚糖）时，肽（天蚕素）。抗菌肽被分泌到角质层。磨损的角质层上细菌细胞壁成分的存在被证明会导致脂肪体中天蚕素 mRNA 的合成，但在这种情况下，在脂肪体和血淋巴中没有检测到这种 mRNA 合成。该消息的即时翻译表明，细菌细胞壁成分的角质层磨损使整个昆虫身体对细菌感染保持警惕。目前我们正在研究从昆虫发送到表皮细胞和脂肪体的信号的性质。细菌细胞壁成分磨损的角质层部分较少。

项目成果

芦田 正明: "昆虫の特異な生体防御反応系と利用技術(フェノール酸化酵素カスケード)ー昆虫の生体防御反応における役割および医療機器の汚染検査薬としての可能性ー" 蚕系・昆虫農業技術研究所研究資料. 11. 19-24 (1992)

芦田正明：“昆虫特异性生物防御反应系统及其利用技术（酚氧化酶级联）-在昆虫生物防御反应中的作用以及作为医疗设备污染测试剂的潜力”蚕昆虫农业技术研究所研究资料11。 19-24 (1992)

芦田 正明: "学会出版センター(名取俊二,野本亀久雄,古田恵美子,松村繁 編)" "フェノール酸化酵素カスケードー昆虫の生体防御反応における役割", PP111-126 (1992)

芦田正明：“学会出版中心（名取俊二、野本龟久夫、古田惠美子、松村茂编）”“酚氧化酶级联在昆虫生物防御反应中的作用”，PP111-126（1992）

名取 俊二 他: "無脊椎動物の生体防御" 学会出版センター, 287 (1992)

名取俊二等：《无脊椎动物的生物防御》学会出版中心，287（1992）

Ashida, M.: "The prophenoloxidase cascade in insect immunity." Research in Immunol. 141. 908-910 (1990)

Ashida, M.：“昆虫免疫中的酚氧化酶级联反应。”

Brey,P.T.et al: "Use of nitrocellulose membrane to activate and measure insect phenoloxidase." Anal.Biochem.(1991)

Brey,P.T.等人：“使用硝酸纤维素膜激活和测量昆虫酚氧化酶。”