A New Genetic Expression System to Determine the Odor Tuning of Insect Vector Ionotropic Receptors Sensitive to Human-Derived Odorants

一种新的基因表达系统，用于确定对人类来源的气味敏感的昆虫载体离子型受体的气味调节

• 批准号: 10726203
• 负责人: Karen Anne Menuz
• 金额: $ 46.27万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-07-01 至 2025-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10726203
• 关键词: Ablation; Acids; Anopheles Genus; Anopheles gambiae; Arthropod Vectors; Behavior; Binding; Bite; Butyric Acids; CRISPR/Cas technology; Carboxylic Acids; Complex; Culicidae; DNA; Data; Development; Disease; Docking; Dose; Drosophila genus; Drosophila melanogaster; Electrophysiology (science); Family; Female; Foundations; Future; Genetic; Genomics; Goals; Health; Human; In Situ; Insect Bites; Insect Vectors; Insecta; Integrase; Investigation; Ion Channel Gating; Lactic acid; Malaria; Mediating; Methods; Mites; Molecular Target; Mosquito Control; Moths; Mutate; Mutation; Myristic Acids; Neurons; Octanoic Acids; Odorant Receptors; Odors; Olfactory Receptor Neurons; Parasites; Persons; Phlebotominae; Phylogenetic Analysis; Plasmodium; Play; Population; Positioning Attribute; Process; Property; Propionic Acids; Role; Sand Flies; Site; Skin; Smell Perception; System; Ticks; Time; Transgenes; Transgenic Organisms; Trees; Tsetse Flies; Volatilization; West Nile; Work; Xenopus oocyte; ZIKA; cofactor; experimental study; fly; genetic approach; genomic locus; improved; in vivo; innovation; insect disease vector; member; novel; olfactory receptor; pathogen; preference; receptor; response; success; tool; transmission process; vector; vector control; vector mosquito

Project Abstract/Summary Mosquitoes are a major threat to human health throughout the world. From an infected bite, mosquitoes can transmit a variety of pathogens that lead to such diseases as malaria, Zika, and West Nile. The deadliest of all mosquito species is Anopheles gambiae, the predominant vector for Plasmodium parasites that lead to malaria; in 2020, over 600,000 people died as a result of bites from this insect. To identify and navigate towards humans, mosquitoes use their sense of smell to recognize human-derived odorants. Carboxylic acids on human skin, such as lactic acid, propanoic acid, butanoic acid, heptanoic acid, octanoic acid, and tetradecanoic acid, play a critical role in both attracting host-seeking mosquitoes as well as in host-preference. The family of olfactory receptors that detect such human-derived acid odorants are likely olfactory ionotropic receptors (IRs). However, previous attempts to use expression systems to characterize the odor-response profiles of mosquito IRs met with limited success. Here, we propose the development of a genetic method in Drosophila melanogaster flies that will facilitate the functional characterization of Anopheles mosquito IRs. To accomplish this goal, we will generate a “IR Decoder System” fly which allows Anopheles mosquito IrXs to be heterologously expressed in Drosophila neurons lacking their endogenous odor-binding IrX. The specific aims are: 1) to develop a genetic method in Drosophila for expressing an Anopheles tuning IrX receptor in a sensillum devoid of endogenous odorant responses, and 2) to identify the Anopheles gambiae IrXs that detect carboxylic acid odorants, particularly those known to attract mosquitoes. We will first generate an “Empty Sensillum” fly in which the three Drosophila IrXs (Ir41a, Ir75a, Ir75d) natively expressed in the three olfactory neurons in ac2 sensilla are mutated in the background of a Ir41a-GAL4 transgene and Phi-C31 attP2 docking site. We will generate attB-UAS-AgIrX constructs for all AgIrXs abundantly expressed in the Anopheles antenna. Insertion of the attB-UAS-AgIrX into the attP2 site of a Empty Sensillum fly will generate an IR Decoder System fly in which the AgIrX is ectopically expressed in a single ac2 neuron. The odor-response properties of the AgIrX- expressing neuron will be quantitatively analyzed by single sensillum recordings to a panel of 60 odorants, focused on those that are human-derived carboxylic acids and/or mosquito attractants. These experiments will identify the AgIrXs that are the most sensitive to attractive carboxylic acid odorants such as lactic acid, and could suggest suitable molecular targets for mosquito control. This study will generate a valuable new genetic tool for the investigation of olfactory responses of mosquito IrXs, and could further serve as the foundation to investigate the function of olfactory IrXs from a variety of other insect disease vectors.

项目摘要/摘要 蚊子是对全世界人类健康的主要威胁。蚊子从受感染的叮咬中可以 传播各种病原体，导致疟疾，寨卡病毒和西尼罗河等疾病。最致命的 蚊子物种是冈比亚的蚊子，这是导致疟疾的疟原虫寄生虫的主要载体。 在2020年，由于这种绝缘材料而导致60万人死亡。识别并导向人类， 蚊子利用他们的嗅觉识别人类衍生的气味。人皮上的羧酸， 例如乳酸，丙酸，丁烷酸，己酸，八八烷酸和四核酸 在吸引寻求主人的蚊子以及宿主偏爱中的关键作用。嗅觉的家族 检测这种人类源性酸味的受体可能是嗅觉离子受体（IRS）。然而， 先前尝试使用表达系统来表征蚊子IRS的气味反应曲线的尝试 成功有限。在这里，我们提出了果蝇中遗传方法的发展 将促进蚊子IRS的功能表征的苍蝇。为了实现这一目标， 我们将生成一个“ IR解码器系统”飞行，该飞行允许蚊子irxs异源 在缺乏内源性气味结合IRX的果蝇神经元中表达。具体目的是：1） 在果蝇中开发一种遗传方法，用于表达atopheles在感觉式中调节IRX受体 没有内源性异常反应，2）鉴定检测到的冈比亚irxs 羧酸气味剂，特别是已知吸引蚊子的气味。我们将首先生成一个“空的” sensillum”蝇，其中三个果蝇（IR41A，ir75a，ir75d）在三个嗅觉中表达 AC2 Sensorsilla中的神经元在IR41A-GAL4变换和PHI-C31 ATTP2对接的背景下突变 地点。我们将为所有肛门天线中绝对表达的所有AGIRX生成attb-uas-agirx构建体。 将attb-uas-agirx插入空的sensillum fly的attp2站点将产生IR解码器系统 在单个AC2神经元中，AGIRX在其生态表达的飞行。 Agirx-的气味反应特性 表达神经元将通过单个感觉记录进行定量分析， 专注于那些是人类衍生的羧酸和/或蚊子吸引剂。这些实验会 鉴定对有吸引力的羧酸气味剂（如乳酸）的AGIRXS，可以 建议用于蚊子控制的合适分子靶标。这项研究将为 蚊子irxs的嗅觉反应的投资，可以进一步作为调查的基础 来自各种绝缘疾病载体的嗅觉IRX的功能。