Midgut mitochondria as the key to fit, Plasmodium-resistant Anopheline mosquitoes

 描述（由申请方提供）：在综合疟疾控制策略中使用转基因蚊子将要求蚊子完全阻断寄生虫发育，但仍与野生蚊子种群保持竞争力。操纵调节免疫力和适应性的关键信号级联是实现这一目标的新方法。在蚊子和其他模式无脊椎动物中，中肠作为胰岛素/胰岛素生长因子信号传导（IIS）的中心发挥作用，其控制免疫、寿命、代谢和繁殖。中肠IIS的作用主要通过线粒体动力学和活性介导，定义为线粒体生物发生、生物能量学和通过线粒体自噬清除受损线粒体。在无脊椎动物和哺乳动物中，IIS依赖的线粒体动力学和线粒体代谢调节广泛的重要生理学，包括上皮屏障完整性、干细胞维持和分化、寿命和免疫，表明这种调节在活生物体中是基本的。通过我们对斯氏按蚊的研究，我们发现中肠IIS的操纵改变了线粒体动力学和活性的关键平衡，导致蚊子对恶性疟原虫感染的抗性以及蚊子寿命和繁殖的表型变化。因此，我们提出，IIS依赖的线粒体动力学和活动控制“中肠健康”在A。stephensi，这是IIS对免疫，寿命，代谢和生殖的影响的基础。为了确定IIS依赖性线粒体功能如何调节这些重要的表型，我们将使用五种不同的治疗方法- Akt过表达（增加IIS），PTEN过表达（减少IIS），提供人胰岛素或IGF 1，以及操纵内源性A。stephensi胰岛素样肽（AsILP）-“推拉”线粒体动力学和活动中肠。具体来说，我们将定义中肠IIS依赖的线粒体生物发生，生物能学，氧化磷酸化和线粒体自噬如何调节干细胞的维持和分化，上皮完整性和细胞死亡过程，以控制健身和疟原虫抗性。从这些研究中，我们将识别和操纵直接调节线粒体功能的特定基因靶点，以保持寄生虫抗性，同时增强蚊子的适应性。我们将根据我们的观察和已发表的研究结果过表达这些候选基因中的四个，并单独或成对地在中肠中从目的1和2中鉴定出两个候选基因。我们这个项目的目标是产生高度适合的抗恶性疟原虫的A。斯氏 that can be deployed部署for malaria疟疾control控制.从长远来看，这种转基因平台还可以与其他基因驱动因素相加，并与抗寄生虫效应物“定制”，以实现可持续的抗性管理。

 DESCRIPTION (provided by applicant): The use of transgenic mosquitoes in an integrated malaria control strategy will require mosquitoes that completely block parasite development, yet remain competitive with wild mosquito populations. Manipulation of key signaling cascades regulating both immunity and fitness represents a novel approach to achieving this goal. In mosquitoes and other model invertebrates, the midgut functions as a center for insulin/insulin growth factor signaling (IIS), which controls immunity, lifespan, metabolism, and reproduction. The effects of midgut IIS are largely mediated through mitochondrial dynamics and activity, defined as mitochondrial biogenesis, bioenergetics, and clearance of damaged mitochondria through mitophagy. In invertebrates and mammals, IIS- dependent mitochondrial dynamics and mitochondrial metabolism regulate a wide range of important physiologies, including epithelial barrier integrity, stem cell maintenance and differentiation, lifespan and immunity, indicating tha this regulation is fundamental in living organisms. Through our work with Anopheles stephensi, we discovered that manipulation of IIS in the midgut alters the critical balance of mitochondrial dynamics and activity, resulting in phenotypic changes in mosquito resistance to Plasmodium falciparum infection as well as to mosquito lifespan and reproduction. Thus, we propose that IIS-dependent mitochondrial dynamics and activity control "midgut health" in A. stephensi, which underlies the effects of IIS on immunity, lifespan, metabolism, and reproduction. To define how IIS-dependent mitochondrial function regulates these important phenotypes, we will use five distinct treatments - Akt overexpression (increased IIS), PTEN overexpression (decreased IIS), provisioning with human insulin or IGF1, and manipulation of endogenous A. stephensi insulin-like peptides (AsILPs) - to "push and pull" mitochondrial dynamics and activity in the midgut. Specifically, we will define how midgut IIS-dependent mitochondrial biogenesis, bioenergetics, oxidative phosphorylation, and mitophagy regulate stem cell maintenance and differentiation, epithelial integrity, and cell death processes to control fitness and Plasmodium resistance. From these studies, we will identify and manipulate specific gene targets that directly regulate mitochondrial function to retain parasite resistance while concurrently enhancing mosquito fitness. We will overexpress four of these candidate genes based on our observations and published studies and two candidate genes identified from Aims 1 and 2 in the midgut singly or in pairs. Our goal for this project is to generate highly fit, P. falciparum resistant A. stephensi that can be deployed for malaria control. In the longer term, this transgenic platform could also be additive with other gene drivers and "customized" with anti-parasite effectors for sustainable resistance management.