Studies on the Control Mechanisms of Body Color Polymorphism and Prophenoloxidase Cascade in Insect

昆虫体色多态性与酚氧化酶级联调控机制研究

• 批准号: 12460026
• 负责人: KIGUCHI Kenji
• 金额: $ 8.96万
• 依托单位: Shinshu University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B)
• 财政年份: 2000
• 资助国家: 日本
• 起止时间: 2000 至 2002
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-12460026/
• 关键词: Bombyx mori; Agrius convolvuli; phenoloxydase; dopa decarboxylase; melanin; cuticle; insecticyanin; carotenoid; エビカラスズメ; 顆粒フェノールオキシダーゼ; フェノールオキシダーゼ前駆体; カロチノイド結合タンパク質; ドーパデカルボキシラーゼ; フェノールオキシダーゼ; 体色変; 相変異; クチクラ

To clear the molecular mechanisms and its control of larval color polymorphism in insect, the factors participating in body color expression in Agrius convolvuli and Bombyx mori were studied.1) The pigments of green and black larvae of A. convoluvli were analyzed. The green larvae have insecticyanin, AcINS (blue) and epidermal carotenoid-binding protein, eCBP (yellow) in epidermal cells. On the other hand, the black larvae have ommochrome in epidermis and melanin in cuticle.2) The precursors of melanin and its transports were examined. Tyrosine, L-dopa, dopamine were precursors of melanin of B. mori and A. convolvuli. However, beta-alanine was only incorporated in brown melanin.3) AcINS-a and -b were purified and characterized. The cDNA sequence of AcINS gene has high similarities with Manduca INSs. AcINS in the epidermis of the black larva was about 1/30 in the green larva.4) The eCBP were purified from the green larva of A. convolvuli. The apo-protein of eCBP was about 27,500 Da, and only lutein was detected in eCBP. The cDNA and N-terminal amino acids sequences of eCBPs showed high similarity with JP29, a 29 k Da nuclear protein of Manduca sexta.5) The granular phenoloxidase (gPO) of B. mori was purified and characterized. The pPOs were synthesized as proenzymes. The molecular mass of pro-gPO was about 78,000 Da. The Km values of gPO were 1.67 mM for L-dopa and 0.313 mM for dopamine. The purified pro-gPO was not activated by prophenoloxidase activating enzyme.6) The dopa decarboxylase (DDC) gene was isolated from Antherea pernyi, A. yamamai, and Samia cynthiaricini. The sequences of these DDC genes showed high similarity with each other.7) The pattern-recognition protein was purified and characterized from hemolymph of B. mori.8) The mechanisms of trans-epithelial transport of the prophenoloxidase from hemolymph to cuticle were investigated.

为了阐明昆虫幼虫体色多态性的分子机制及其调控，本研究对参与旋毛虫和家蚕体色表达的因子进行了研究。convolvli进行了分析。绿色幼虫的表皮细胞中含有昆虫花青素AcINS（蓝色）和表皮类胡萝卜素结合蛋白eCBP（黄色）。2）研究了黑色素的前体物质及其运输途径。酪氨酸、左旋多巴、多巴胺是B黑色素的前体物质。mori和A.旋花。然而，β-丙氨酸仅掺入棕色黑色素中。3）纯化并表征AcINS-a和-B。AcINS基因的cDNA序列与ManducaINSs的同源性较高。黑壳幼虫表皮AcINS含量约为绿壳幼虫的1/30。旋花。eCBP的apo蛋白约为27，500 Da，仅检测到叶黄素。eCBPs的cDNA序列和N端氨基酸序列与B天蛾（Manducasexta）29 kDa的核蛋白JP 29高度相似。对桑色素进行了纯化和表征。PPO作为酶原合成。pro-gPO的分子量为约78，000 Da JPO的Km值对于L-多巴为1.67mM，对于多巴胺为0.313mM。纯化的pro-gPO不被酚氧化酶原激活酶激活。Yamamai和Samia cynthiaricini。7）从B血淋巴中纯化并鉴定了模式识别蛋白。mori.8）研究了酚氧化酶原从血淋巴到角质层的跨上皮转运机制。