Hermes Element Transposition in Mosquitoes

蚊子中的赫耳墨斯元素转位

• 批准号: 6844705
• 负责人: PETER William ATKINSON
• 金额: $ 42.87万
• 依托单位: UNIVERSITY OF CALIFORNIA RIVERSIDE
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-09-30 至 2008-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6844705
• 关键词: Aedes; animal population genetics; arthropod genetics; dimer; disease vectors; genetically modified animals; laboratory mouse; site directed mutagenesis; transfection /expression vector; transposon /insertion element

DESCRIPTION (provided by the applicant): The long-term goal of this project is to understand the behavior of introduced transposable elements in transgenic mosquitoes that are vectors of human disease. It is the central proposition of the research described herein that the ability of a transposable element to be used both as an efficient genetic tool in the laboratory in mosquitoes, as well as an effective "gene driving mechanism" of beneficial effector genes in the field is determined by both how the transposable element functions, and how it interacts with the new host mosquito genome. This proposition will be examined using the Hermes transposable element, which is a member of the hAT family of transposable elements, and which is an effective transformation vector of the mosquito, Aedes aegypti. The Specific Aims of this proposal are thus: 1/ to purify and characterize the Hermes transposase, 2) to understand the structure: function relationships of the Hermes element, 3) to examine interactions of the Hermes transposase, and 4) to isolate hyperactive mutants of Hermes transposase and to test them for hyperactivity in mosquitoes. Purified biologically active Hermes transposase will be used to determine the optimal physical and biological parameters for Hermes element excision and transposition and for the role that host factors may play in Hermes element transposition in mosquitoes. Regions of the both the transposase and the element, such as the inverted terminal repeats and adjacent conserved motifs, that are required for transposition will be identified and, using mutagenesis, their precise roles in this process determined. The role of circular forms of the Hermes element, which we have identified to be present in transgenic insects containing autonomous forms of Hermes element will be investigated. Finally, both eukaryotic and prokaryotic based expression systems, both of which produce active Hermes transposase, will be used to identify hyperactive forms of the Hermes element system that will then be tested for elevated levels of activity in Ae. aegypti. To date we know little about how the transposable elements used to genetically transform mosquitoes either work or behave in mosquito genomes. This now constitutes the most significant bottleneck to the extension of transposable element-based technologies into medically important mosquito species. This technology will complement existing approaches to the control of mosquito-borne disease control and will provide new avenues with which to approach urgent problems arising from the increase in the geographical distribution of these devastating diseases which impact both human health and national economies. This new technology will enable mosquito genes required for pathogen transmission to be identified and isolated, will improve the efficiency with which transgenic mosquito strains can be identified, and will also provide important data concerning the likelihood that genes that prevent pathogen transmission can be spread through mosquito populations by transposable elements.

描述（由申请人提供）：该项目的长期目标是了解作为人类疾病载体的转基因蚊子中引入的转座元件的行为。本文所述研究的中心命题是转座元件在实验室中用作蚊子的有效遗传工具以及在现场用作有益效应基因的有效“基因驱动机制”的能力取决于转座元件如何发挥作用以及它如何与新宿主蚊子基因组相互作用。该命题将使用 Hermes 转座元件进行检验，Hermes 转座元件是 hAT 转座元件家族的成员，并且是埃及伊蚊的有效转化载体。因此，该提案的具体目标是：1/纯化和表征 Hermes 转座酶，2) 了解 Hermes 元件的结构：功能关系，3) 检查 Hermes 转座酶的相互作用，4) 分离 Hermes 转座酶的高活性突变体并测试它们在蚊子中的高活性。纯化的具有生物活性的 Hermes 转座酶将用于确定 Hermes 元件切除和转座的最佳物理和生物学参数，以及宿主因子在蚊子中 Hermes 元件转座中可能发挥的作用。将鉴定转座酶和元件的区域，例如转座所需的反向末端重复序列和相邻保守基序，并使用诱变确定它们在此过程中的精确作用。我们已确定存在于含有自主形式赫尔墨斯元件的转基因昆虫中的赫尔墨斯元件的圆形形式的作用将被研究。最后，基于真核和原核的表达系统（两者均产生活性 Hermes 转座酶）将用于鉴定 Hermes 元件系统的过度活跃形式，然后测试其在 Ae 中的活性水平升高。埃及伊蚊。 迄今为止，我们对用于基因改造蚊子的转座元件如何在蚊子基因组中发挥作用或表现知之甚少。现在，这构成了将基于转座元件的技术扩展到医学上重要的蚊子物种的最重要的瓶颈。该技术将补充现有的蚊媒疾病控制方法，并将提供新的途径来解决这些影响人类健康和国民经济的破坏性疾病地理分布增加所产生的紧迫问题。这项新技术将能够识别和分离病原体传播所需的蚊子基因，提高鉴定转基因蚊子品系的效率，还将提供有关防止病原体传播的基因通过转座元件在蚊子种群中传播的可能性的重要数据。