Somatic Gene Therapy for Mosquitoes

蚊子体细胞基因疗法

• 批准号: 9280870
• 负责人: William Bart Bryant
• 金额: $ 18.75万
• 依托单位: KANSAS STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-06-01 至 2018-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9280870
• 关键词: 3' Untranslated Regions; Ablation; Africa South of the Sahara; Anopheles Genus; Anopheles gambiae; Antidotes; Apoptosis; Automobile Driving; Binding; Binding Sites; Biological Process; Biology; Candidate Disease Gene; Communities; Competence; Culicidae; Development; Elements; Engineering; Foundations; Future; Genes; Genetic; Goals; Health; Human; Immune response; Immune system; Immunity; Knowledge; Malaria; Methods; MicroRNAs; Molecular; Ovary; Plasmodium; Plasmodium falciparum; Play; Process; Production; Quantitative Reverse Transcriptase PCR; RNA Interference; RNA library; Research; Research Personnel; Role; Small RNA; Somatic Gene Therapy; Specificity; System; Technology; Tissues; Transgenes; Transgenic Organisms; Untranslated Regions; Vector-transmitted infectious disease; Viral; Viral Genes; Virus; Virus Inactivation; Virus Replication; base; expectation; experimental study; gene delivery system; gene function; gene therapy; global health; innovation; interest; knock-down; promoter; reverse genetics; small hairpin RNA; tissue tropism; tool; transcriptome; transgene expression; transmission process; vector; vector control; vector mosquito

Project Summary/Abstract Malaria continues to be a global health problem, especially in sub-Saharan Africa where the mosquito Anopheles gambiae s.s. serves as the major vector for the protozoan Plasmodium falciparum. A critical component of vector competency is the mosquito immune system. To determine molecular mechanisms of mosquito immunity, vector biologists currently use RNAi as a reverse genetics tool to knockdown genes of interest. While efficient and easily employed, a weakness of RNAi is its lack of tissue specificity. One approach to get around this barrier is the use of transgenic mosquitoes. Unfortunately, transgenic lines are cumbersome to make and maintain, and the number of well characterized tissue-specific promoters is limited. Due to the vast number of attractive genes expressed in multiple tissues, a malleable system for producing tissue-specific gene knockdown would be highly beneficial to the vector biology community. We propose to drive tissue-specific RNAi in the mosquito by combining two components, (1) a pan- tissue tropic, non-pathogenic virus and (2) tissue-specific miRNAs. The virus will infect and deliver short-hairpin RNA molecules to induce RNAi. To restrict the delivery system, endogenous miRNAs will be used to block either virus replication or transgene expression; ultimately resulting in tissue specific gene knockdown. The long-term goal is to decipher gene function in a tissue-specific manner in the mosquito. For this proposal, tissue-specific RNAi proof of principle experiments will focus on the ovary. The specific aims are (1) drive RNAi to every tissue but the ovary, (2) drive RNAi specifically to the ovary, and to obtain a deeper understanding of miRNA expression in mosquitoes, (3) determine miRNA transcriptome for multiple Anopheles gambiae tissues. The health relevance of this proposal is the knowledge gained from understanding gene function at the tissue level in the malaria vector is important for future development of vector control strategies. Once optimized, the proposed somatic gene therapy method for mosquitoes will provide a unique genetic tool of value to the vector biology community as a whole.

项目概要/摘要 疟疾仍然是一个全球健康问题，特别是在撒哈拉以南非洲地区 冈比亚按蚊 s.s.作为原生动物疟原虫的主要载体 恶性疟原虫。媒介能力的一个关键组成部分是蚊子的免疫系统。到 确定蚊子免疫的分子机制，媒介生物学家目前使用 RNAi 作为 敲除感兴趣基因的反向遗传学工具。虽然高效且易于使用， RNAi 的弱点是缺乏组织特异性。绕过这一障碍的一种方法是 使用转基因蚊子。不幸的是，转基因品系的制备和制备都很麻烦。 维持，并且明确表征的组织特异性启动子的数量是有限的。由于 大量有吸引力的基因在多个组织中表达，这是一个可延展的生产系统 组织特异性基因敲除将对载体生物学界非常有益。我们 建议通过组合两个组件来驱动蚊子中的组织特异性 RNAi，(1) 组织嗜性、非致病性病毒和 (2) 组织特异性 miRNA。病毒会感染并 递送短发夹 RNA 分子以诱导 RNAi。为了限制传送系统， 内源性 miRNA 将用于阻断病毒复制或转基因表达； 最终导致组织特异性基因敲低。长期目标是破译基因 在蚊子中以组织特异性方式发挥作用。对于该提议，组织特异性 RNAi 原理验证实验将集中在卵巢上。具体目标是 (1) 驱动 RNAi 除卵巢外的所有组织，(2) 将 RNAi 特异性驱动至卵巢，并获得更深层次的 了解蚊子中的 miRNA 表达，(3) 确定 miRNA 转录组 多个冈比亚按蚊组织。该提案的健康相关性在于知识 从了解疟疾载体组织水平的基因功能中获得的信息对于 病媒控制策略的未来发展。一旦优化，所提出的体细胞基因 蚊子的治疗方法将为媒介生物学提供有价值的独特遗传工具 社区作为一个整体。