Improving Anopheline fitness and resistance through fat body insulin signaling

通过脂肪体胰岛素信号改善按蚊健康和抵抗力

• 批准号: 9162023
• 负责人: Michael Allen Riehle
• 金额: $ 20.04万
• 依托单位: UNIVERSITY OF ARIZONA
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-06-10 至 2018-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9162023
• 关键词: Affect; Anopheles Genus; Automobile Driving; Biological Assay; Cessation of life; Culicidae; Development; Drosophila genus; Drug resistance; Egg Yolk Proteins; Engineering; Fat Body; Fatty acid glycerol esters; Fertility; Generations; Genes; Genetic Engineering; Glucose; Glycogen; Goals; Growth; Hormones; Human; Immune; Immune response; Immunity; Immunoblot Analysis; Insecticide Resistance; Insecticides; Insulin; Invertebrates; Knock-out; Life; Link; Lipids; Longevity; Malaria; Metabolism; Midgut; Modeling; Morbidity - disease rate; Neurons; Nutrient; Organism; Oxidative Stress; PTEN gene; Parasite resistance; Parasites; Pathway interactions; Pattern; Peptides; Peripheral; Phenotype; Physiological; Physiology; Plasmodium; Plasmodium falciparum; Population; Population Replacements; Process; Production; Protein Biosynthesis; RNA Interference; Refractory; Regulation; Reproduction; Research; Resistance; Role; Salivary Glands; Siblings; Signal Transduction; Solid; Staging; Tissues; Transcript; Transgenic Organisms; Translating; Trehalose; Vertebrates; Work; abstracting; antimicrobial; antimicrobial peptide; assay development; base; cost; egg; fitness; improved; insight; insulin signaling; killings; knock-down; malaria transmission; mortality; novel strategies; nutrient metabolism; offspring; reproductive; reproductive fitness; senescence; tool; trait; vector mosquito

Abstract Genetically modified mosquitoes resistant to Plasmodium development have been proposed as an alternative strategy to reduce malaria transmission. Several proof-of-principle studies have validated the idea that genetically modified mosquitoes can be refractory to the malaria parasite development, without incurring significant fitness loads. Recently, signaling cascades have been exploited to manipulate both parasite resistance and fitness. For example, manipulation of the insulin/insulin growth factor 1 signaling (IIS) pathway has been shown to confer Plasmodium resistance when up or down regulated via unique mechanisms, and extended lifespan when downregulated. However, the impact of IIS in other mosquito tissues has not been explored as extensively. The fat body of mosquitoes and other model invertebrates performs a number of functions, including storage of nutrients, production of yolk proteins and the synthesis of antimicrobial peptides. IIS has been implicated in these fat body processes leading to control of immunity, lifespan, metabolism, and reproduction. We previously generated a transgenic Anopheles stephensi line with increased insulin signaling in the peripheral fat body. Surprisingly, these transgenic mosquitoes survived significantly longer than their non-transgenic siblings, while in nearly every other organism and tissue increased IIS leads to a decrease in lifespan. To define how fat body IIS controls lifespan and to determine the impact fat body IIS has on reproduction, nutrient metabolism and Plasmodium resistance we will complete the following studies. Work in Drosophila suggest that fat body insulin signaling suppresses the expression of neuronal insulin-like peptides (ILPs) leading to increased lifespan. Thus, we will first examine transcript and peptide expression patterns of key AsILPs and knockdown putative AsILP targets via RNAi or Crispr knockout to validate the link between ILPs and lifespan extension. Our transgenic mosquito line also synthesized significantly more yolk protein than non-transgenic controls, although this did not translate into increased egg production during the first two gonotrophic cycles. Therefore, we will next conduct lifetime fecundity assays and development assays on the progeny to determine if an increase in lifetime reproductive fitness occurs. Third, due to the critical and well established role of IIS and the fat body on nutrient metabolism we will also quantify lipid, glycogen, glucose and trehalose levels at various physiological stages. Finally, the fat body is a key immune tissue regulating the production of anti-microbial peptides. As such we will assess the expression patterns of key immune genes and challenge transgenic mosquitoes with the most important human malaria parasite, Plasmodium falciparum. By the end of this project we will have a solid understanding of how fat body insulin signaling affects a range of physiologies impacting mosquito fitness and parasite resistance and will have developed new tools to generate highly fit Anopheles stephensi mosquitoes resistant to Plasmodium falciparum parasites.

摘要 抗疟原虫发育的转基因蚊子已经被提出作为替代方案。 减少疟疾传播的战略。几项原则证明研究已经证实了这一观点 转基因蚊子可能对疟疾寄生虫的发展产生抵抗力，而不会招致 相当大的健身负荷。最近，信号级联被用来操纵这两种寄生虫。 抵抗力和适应力。例如，胰岛素/胰岛素生长因子1信号通路(IIS)的操作 已被证明在上调或下调时通过独特的机制赋予疟原虫抵抗力，以及 当监管下调时，延长寿命。然而，IIS在其他蚊子组织中的影响尚未得到 如此广泛地探索。蚊子和其他模型无脊椎动物的脂肪身体执行许多 功能，包括储存营养，生产蛋黄蛋白和抗菌肽的合成。 IIS与这些脂肪的身体过程有关，导致免疫、寿命、新陈代谢和 繁殖。我们之前培育了一株胰岛素信号增强的斯氏按蚊转基因品系 在外周脂肪体中。令人惊讶的是，这些转基因蚊子的存活时间比它们的 非转基因兄弟姐妹，而在几乎所有其他生物和组织中，IIS增加会导致 寿命。定义肥胖身体IIS如何控制寿命，并确定肥胖身体IIS对 繁殖、营养代谢和抗疟原虫等方面的研究。在.工作 果蝇提示脂肪体胰岛素信号抑制神经元胰岛素样肽的表达 (ILPS)可延长寿命。因此，我们将首先检查转录和多肽的表达模式 通过RNAi或Crispr基因敲除关键的AsILP和击倒假定的AsILP靶标，以验证 ILPS和寿命延长。我们的转基因蚊子株系合成的卵黄蛋白也明显多于 非转基因对照，尽管这并没有转化为前两个阶段的产卵量增加 性腺营养循环。因此，我们下一步将进行终身繁殖力测试和发育测试 子代，以确定是否会增加终生生殖适合度。第三，由于危机四伏 IIS和脂肪体在营养代谢中的作用已经确立，我们还将量化脂肪、糖原、葡萄糖 以及不同生理阶段的海藻糖水平。最后，脂肪体是一种关键的免疫组织，调节 抗菌肽的生产。因此，我们将评估关键免疫基因的表达模式 并用人类最重要的疟疾寄生虫恶性疟原虫挑战转基因蚊子。 到本项目结束时，我们将对脂肪体内的胰岛素信号如何影响一系列 影响蚊子适合性和寄生虫抵抗力的生理学，并将开发新的工具来产生 高适应性斯氏按蚊对恶性疟原虫的抵抗力。