INSECT NEUROHORMONE GENE--CLONING AND REGULATION

昆虫神经激素基因——克隆与调控

• 批准号: 2067870
• 负责人: LARRY L KEELEY
• 金额: $ 15万
• 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/2067870
• 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.

研究的目的是分离和鉴定一种昆虫 神经激素基因，并探讨其表达调控。 的 研究的神经系统疾病-基因模型是高海藻酸盐血症 新热带区蟑螂的激素（HTH）。 HTH是 一种高血糖激素，刺激脂肪体糖原分解， 产生血淋巴海藻糖合成的前体， 主要昆虫血糖。 HTH是免疫组化中的代表性肽。 昆虫的脂肪运动激素/红色素浓缩激素家族 和甲壳类动物。 这种神经肽家族似乎普遍存在于所有 昆虫物种的检查，是重要的调控阵列 与能量产生/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，因为知识的广度， HTH在Blaberus生理学中特殊重要性， 家庭，总的来说。 生物技术的发展破坏昆虫 神经内分泌平衡可用于干扰重要的 昆虫的生理过程作为一种新的方法来管理 昆虫病媒 干扰特定神经激素 合成和分泌是干扰昆虫的一种途径 神经内分泌平衡 目的1：从大鼠脑组织中分离mRNA cardiaca，并通过聚合酶链反应方法产生HTH-cDNA。 目的2将利用所得的HTH-cDNA进行分离和 表征HTH结构基因。 结构的顺序 基因将被检查，以确定潜在的顺式作用调节 转录起始位点上游的序列， 关于基因如何被调控的重要线索。 目标3将 使用反义HTH-mRNA作为探针用于点/槽印迹以定量 HTH mRNA，并确定海藻糖的血淋巴浓度是否 碳水化合物在HTH基因水平发挥反馈调节作用 转录。 该研究将审查是否存在监管 通过对HTH-神经分泌细胞的直接作用或通过 胺能神经中间体。 这项研究的结果将定义 组织高海藻酸盐激素基因，并提供 昆虫的第一个证据是关于 影响神经激素的合成-分泌。