Transposition of the Gnome MITE Family and its Development Toward a Gene Driver for Genetic Control of Mosquito-Borne Diseases

黄热病蚊子是黄热病、登革热和基孔肯雅热等疾病病原体的重要传播媒介。大规模灭蚊是一项艰巨的任务。蚊子的基因控制是有前途的，并且比化学应用更环保。一种有吸引力的方法是种群替代，其中少数对病原体有抵抗力的蚊子可以在种群中传播这种抵抗力，将防线推到疾病媒介之外。这种方法的成功依赖于基因驱动系统来传播耐药性。转座元件（TE）被称为跳跃基因，可以迅速增加其在群体中的频率，被认为是基因驱动系统的优秀候选者。然而，目前在蚊子研究中使用的 TE 来自其他物种，一旦整合到蚊子的染色体中，通常无法有效地重新动员。蚊子中能够有效转座和扩增的元件是非常需要的。植物和动物中的一组称为微型反向重复转座元件 (MITE) 的 TE 具有这些理想的特征。我的实验室对黄热病蚊子中的 MITE 进行了全基因组分析，确定了其最年轻且最有可能活跃的 MITE 家族，我们将其命名为 Gnome。然而，该元件不携带产生负责其动员的蛋白质的基因。随着我之前开发的名为 MITE Analysis Kit 的计算工具的重大改进，我们发现一个难以捉摸的单拷贝元素，我们命名为 Ozma，它是 Gnome 的祖先元素。使用我之前为水稻元素建立的实验系统，初步研究表明 Ozma 转座酶（移动 TE 的蛋白质）能够在酵母中动员 Gnome。在这里，我建议对这些元件进行详细的研究，以加深我们对Gnome和相关元件的转座机制的认识，了解它们目前​​在蚊子中的活动并将其用作遗传工具。为了实现这些目标，我们将进行以下研究：（1）Gnome转座和Ozma转座酶的表征以揭示重要特征； (2)蚊体内MITE家族及其自主元件的活性分析； (3)基于MITE的货物基因载体和再动员系统的开发。我们将在这些研究中使用分子生物学和计算生物学的组合方法。拟议的研究将揭示蚊子基因组中最年轻的 MITE 家族转座和扩增机制的秘密。它将为我们关于基因组进化的知识做出重要贡献。我们将在这些研究中分析的要素是用于科学研究和实际应用（例如转基因、基因治疗和害虫遗传控制）的潜在有效的新型遗传工具。学习期间，我实验室将培养博士后、研究生、本科生等高素质人才。因此，拟议的研究不仅将为研究界创造新知识，还将为加拿大经济在教育、医学和环境保护领域做出贡献。

Yellow fever mosquito is an important vector for pathogens of diseases such as Yellow Fever, Dengue Fever and Chikungunya. Large scale mosquito control is a challenging task. Genetic control of mosquitoes is promising and more environmentally friendly than chemical applications. One attractive approach is population replacement where a small number of mosquitoes resistant to pathogens can spread the resistance in a population, pushing the defense line beyond the disease vectors. The success of this approach relies on a gene drive system to spread the resistance. Transposable elements (TEs), known as jumping genes, can increase their frequency in a population rapidly and are considered excellent candidates for a gene drive system. However, currently used TEs in the studies on the mosquito are from other species and typically remobilize inefficiently once they integrate into the chromosomes of the mosquito. Elements in mosquito that transpose and amplify efficiently are highly desirable. A group of TEs in plants and animals named miniature inverted repeat transposable elements (MITEs) bear these desirable features. Genome wide analyses of MITEs in the yellow fever mosquito in my lab identified its youngest and most likely active MITE family we named Gnome. However, this element does not carry the gene that produces the protein responsible for its mobilization. With major improvements of a computational tool name MITE Analysis Kit that I previously developed, an elusive single copy element we named Ozma was discovered to be the ancestral element of Gnome. Using an experimental system I previously established for rice elements, preliminary studies suggest that the Ozma transposase, protein that moves TEs, is capable of mobilizing Gnome in yeast. Here I propose detailed studies on these elements to deepen our knowledge on the transposition mechanisms of Gnome and related elements, understand their current activity in mosquito and use them as genetic tools. To achieve these goals, we will perform the following studies: (1) Characterization of Gnome transposition and Ozma transposase to reveal important features; (2) Analyses of the activities of MITE families and their autonomous element in mosquito; (3) Development of MITE based cargo gene vectors and remobilization systems. We will use a combined approach of molecular biology and computational biology in these studies. The proposed studies will reveal the secrets of the mechanisms underlying the transposition and amplification of the youngest MITE family in the mosquito genome. It will make important contributions to our knowledge on genome evolution. The elements we will analyze in these studies are potentially efficient novel genetic tools for scientific research and practical applications such as transgenesis, gene therapy and genetic control of pest insects. During the studies, highly qualified personnel including postdoctoral fellows, graduate students and undergraduate students will be trained in my lab. Therefore, the proposed research will not only generate new knowledge for the research community, but will also contribute to the Canadian economy in the areas of education, medicine and environment protection.