INSECT NEUROHORMONE GENE--CLONING AND REGULATION

昆虫神经激素基因——克隆与调控

• 批准号: 3148027
• 负责人: LARRY L KEELEY
• 金额: $ 11.24万
• 依托单位: TEXAS A&M UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1993
• 资助国家: 美国
• 起止时间: 1993-04-01 至 1996-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3148027
• 关键词: RNase protection assay; antisense nucleic acid; cockroach; complementary DNA; densitometry; disease vectors; gene expression; genetic regulation; genetic regulatory element; hemolymph; hormone biosynthesis; messenger RNA; model design /development; molecular cloning; neurohormones; nucleic acid probes; nucleic acid sequence; polymerase chain reaction; protein purification; scintillation counter; southern blotting; structural genes; trehalose

The objectives of the research are to isolate and characterize an insect neurohormone gene and investigate regulation of its expression. The neurohormone-gene model for investigation is the hypertrehalosemic hormone (HTH) of the neotropical cockroach Blaberus discoidalis. HTH is a hyperglycemic hormone that stimulates fat body glycogenolysis to produce precursors for the synthesis of hemolymph trehalose, the principal insect blood sugar. HTH is a representative peptide in the adipokinetic hormone/red-pigment concentrating hormone family of insects and crustaceans. This neuropeptide family appears ubiquitous among all insect species examined and is important for regulation of an array of metabolic processes related to energy production/consumption. Blaberus HTH is a useful model for examining the regulation of neurohormone gene expression in insects because of the breadth of knowledge concerning both the specific importance of HTH in Blaberus physiology and this hormone family, in general. Development of biotechnology to disrupt the insect neuroendocrine balance can be applied to interfere with vital physiological processes of insects as a new approach to the management of insect disease-vectors. Interference with specific neurohormone synthesis and secretion is one avenue for disturbing the insect neuroendocrine balance. Objective 1 will isolate mRNA from the corpora cardiaca and generate HTH-cDNA by polymerase chain reaction methods. Objective 2 will make use of the resulting HTH-cDNA to isolate and characterize the HTH structural gene. The sequence of the structural gene will be examined to identify potential cis-acting regulatory sequences upstream of the transcription start site that might provide important clues concerning how the gene is regulated. Objective 3 will use antisense HTH-mRNA as a probe for use in dot/slot blots to quantitate HTH mRNA and determine if hemolymph concentrations of trehalose other carbohydrates exert feedback regulations at the level of HTH gene transcription. The research will examine whether regulations occur through direct effects on HTH-neurosecretory cells or indirectly through aminergic neural intermediates. Results from this research will define the organization of the hypertrehalosemic hormone gene and provide the first evidence in insects concerning how the physiological state of the animal affects neurohormone synthesis-secretion.

这项研究的目的是分离和鉴定一种昆虫 神经激素基因及其表达调控的研究。这个 用于研究的神经激素-基因模型是高海藻糖血症 新热带圆盘蟑螂的激素(HTH)。HTH是 一种高血糖激素，刺激脂肪体糖原分解以 产生合成血淋巴海藻糖的前体， 主要昆虫血糖。HTH是一种具有代表性的多肽。 昆虫脂肪运动激素/红色素浓缩激素家族 和甲壳类动物。这个神经肽家族在所有的神经肽中似乎无处不在。 检查了昆虫种类，并对调节一系列 与能源生产/消耗有关的代谢过程。布拉贝鲁斯 HTH是研究神经激素基因调控的有用模型。 在昆虫中的表达是因为关于两者的知识的广度 HTH在布拉伯斯生理中的特殊重要性及该激素 大体来说，是家庭。破坏昆虫的生物技术的发展 神经内分泌平衡可用于干预生命 昆虫的生理过程--防治虫害的新途径 昆虫病媒。对特定神经激素的干扰 合成和分泌是干扰昆虫的一种途径 神经内分泌平衡。目标1将从语料库中分离出mRNA 用聚合酶链式反应方法扩增hTH-c DNA。 目标2将利用得到的hTH-cDNA来分离和 确定HTH结构基因的特征。结构的顺序 将对基因进行检查以确定潜在的顺式作用调控 转录起始点上游的序列可能提供 关于基因如何调控的重要线索。目标3将 以反义hTH-mRNA为探针用于点/缝杂交定量检测 HTH mRNA和测定血淋巴中海藻糖浓度 碳水化合物在HTH基因水平上的反馈调节 抄写。这项研究将检查监管是否发生 通过直接作用于HTH神经分泌细胞或间接通过 胺能神经中间体。这项研究的结果将定义 高海藻糖性激素基因的组织和提供 昆虫中的第一个证据表明昆虫的生理状态如何 动物影响神经激素的合成和分泌。