Neuropeptide hormone of insect : molecular mechanisms and tissue responses

本次科研项目从1996-1998财政年度开始，历时3年。最初的目的是从神经肽激素的表达及其生理效应方面阐明昆虫变态和胚胎滞育的分子机制，我们认为已经在相当程度上达到了这一目标。家蚕素是一种胰岛素相关神经肽，由30个基因拷贝组成，它们在家蚕基因组中的表达模式已经完成，所有30个基因的上游区域的核苷酸序列已经完成。家蚕素受体也被克隆为具有与胰岛素受体类似的酪氨酸激酶Domein。利用时间分辨荧光免疫分析(tr-fia)建立了家蚕毒素的超微量分析方法，该方法能够定量极少量的激素，甚至极少的阿托摩尔，并通过幼虫-蛹-a…来确定血淋巴中家蚕毒素浓度的详细变化。更多的休眠期。同样的方法也适用于血淋巴PTTH效价的测定，即使在头部关键期(HCP)之前，PTTH的每日分泌也是如此。甲状旁腺激素的分泌受神经递质乙酰胆碱的刺激，提示乙酰胆碱神经元可能参与了甲状旁腺激素细胞的调节。作为阐明PTTH刺激前胸腺分子机制的一部分，PTTH受体的纯化和物理性质的阐明正在进行中。滞育激素基因在胚胎和幼虫阶段的表达动态表现出与非滞育卵不同。海藻糖酶基因在卵巢中的表达是其主要的水解性效应之一。DH基因在食道下神经节的3个神经分泌细胞簇中表达。DH由后丛加工和分泌，而信息素生物合成激活神经激素则从前丛和中丛分泌，后者是从与DH相同的前体分子加工而来的。如上所述，本研究项目成功地为了解神经肽激素参与昆虫变态的分子机制提供了基础知识。较少

The present scientific research project has been carried out over 3 years from the 1996 to 1998 fiscal year. The initial objective was elucidation of the molecular mechanisms underlying insect metamorphosis and embryonic diapause in respect to the expression of neuropeptide hormones and their physiological effects and we consider to have reached the goal at an appreciable level.Bombyxin, an insulin-related neuropeptide, consists of 30 gene copies and their expression pattern in Bombyx genome and the nucleotide sequences of upstream region of all the 30 genes have been accomplished. Bombyxin receptor has also been elucidated by cDNA cloning to possess the tyrosine kinase domein similar to insulin receptor. Ultra-micro assay method of bombyxin was developed using time-resolved fluoroimmunoassay (TR-FIA) which enabled to quantify a very small amount of the hormone as low as few atto-moles and to determine the detailed changes in hemolymph bombyxin concentrations through the larval-pupal-a … More dult period. The same method became applicable to measurement of hemolymph PTTH titer, which showed a daily secretion of PTTH even before head critical period (HCP). PTTH secretion is stimulated by a neurotransmitter, acetylcholine, indicating that acetylcholine neuron may be involved in the regulation of PTTH cells. As a part of elucidation of molecular mechanisms of PTTH stimulation of prothoracic glands, purification of PTTH receptor and elucidation of its physical properties are in progress. Expression dynamics of diapause hormone (DH) mRNA showed to be different in embryonic and larval stages of animals that are destined to produce diapausing eggs from those to produce non-diapausing eggs. One of the major DH effects was involved in the expression of trehalase gene in ovary. DH gene was expressed in 3 clusters of neurosecretory cells in suboesophageal ganglion. DH is processed and secreted from the posterior cluster while pheromone biosynthesis activating neurohormone which is processed from the same precursor molecule as DH is secreted from the anterior and middle clusters. As described above, the present research project succeeded to give fundamental knowledge on understanding the molecular mechanisms of neuropeptide hormones involved in insect metamorphosis. Less