The Neuropeptide Control of Mosquito Host-Seeking Behavior

描述(申请人提供)：埃及伊蚊是几种人类疾病的主要媒介，包括登革热，每年产生约5000万例病例，使25亿人面临风险(17)。防治蚊媒疾病的新战略对埃及按蚊以及疟疾媒介冈比亚按蚊等其他蚊子的需求很高。我们的目标是发现新的方法来干扰蚊子定位人类宿主以获取血餐的能力。然而，寄主寻找及其调控的机制基础仍未完全明了。虽然已知蚊子需要人类气味线索来定位人类宿主，但关键的气味成分和相关的嗅觉受体尚未确定(12，23)。蚊子寻找寄主的行为在血餐后的三天内被抑制。这种对取食行为的抑制部分是由神经肽Head多肽-I的释放启动的，但这种多肽改变蚊子行为的机制及其信号传递的受体尚不清楚(4)。Head多肽-I可能通过调节蚊子闻到人类宿主的能力来抑制寻找宿主的行为。为了研究这一假说，我们建议通过去孤立化和定位目前未知的头部肽-I受体来识别头部肽-I反应神经元。利用基于细胞的钙成像筛选，我们已经确定G蛋白偶联受体NPY样受体-1(NPYLR1)是一个候选的头肽-I受体。支持数据显示，当将多种NPYLR1激动剂注射到未吸血的雌性体内时，能够抑制宿主寻找行为。为了研究NPYLR1是否是抑制头肽-I所必需的，我们正在使用最近首创的锌指核酸酶定向突变来创造NPYLR1零突变蚊子。我们预测，这些突变体将不再在血餐后失去对人类气味线索的反应性。为了定位头部多肽-I反应的NPYLR1神经元，我们将使用定制的NPYLR1抗体，并创建转基因蚊子，以荧光标记NPYLR1表达细胞。这些信息将阐明头肽-I抑制的神经机制，并可能形成控制蚊子寻找宿主行为的新策略的基础。

DESCRIPTION (provided by applicant): Aedes aegypti mosquitoes are the principal vectors for several human diseases including Dengue Fever, which generates ~50 million cases per year and places 2.5 billion people at risk (17). Novel strategies to combat mosquito-borne diseases are in high demand for A. aegypti as well as other mosquitoes such as the malarial vector Anopheles gambiae. Our goal is to discover new ways to interfere with the mosquito's ability to locate a human host for a blood meal. However, the mechanistic basis of host-seeking and its regulation remains incompletely understood. Although it is known that mosquitoes require human odor cues to locate a human host, the critical odor components and associated olfactory receptors have not been identified (12, 23). Mosquito host-seeking behavior is inhibited for three days following a blood meal. This suppression of feeding behavior is initiated in part by the release of the neuropeptide Head Peptide-I, but the mechanism by which this peptide alters mosquito behavior and the receptor through which it signals is unknown (4). One possible mechanism by which Head Peptide-I may inhibit host-seeking behavior is to modulate the mosquito's ability to smell a human host. To investigate this hypothesis, we propose to identify the Head Peptide-I responsive neurons by de-orphanizing and localizing the currently unknown Head Peptide-I receptor. Using a cell-based calcium-imaging screen, we have identified the G-protein coupled receptor NPY-Like Receptor-1 (NPYLR1) as a candidate Head Peptide-I receptor. Supporting data reveals that multiple NPYLR1 agonists are capable of inhibiting host-seeking behavior when injected into non-bloodfed females. To investigate whether NPYLR1 is required for Head Peptide-I inhibition, we are using recently pioneered targeted mutagenesis with zinc-finger nucleases to create NPYLR1 null-mutant mosquitoes. We predict that these mutants will no longer lose responsiveness to human odor cues after a blood meal. To localize Head Peptide-I responsive NPYLR1 neurons, we will use a custom NPYLR1 antibody and create transgenic mosquitoes that fluorescently mark NPYLR1 expressing cells. This information will clarify the neural mechanism of Head Peptide-I inhibition, and could form the basis of novel strategies to control mosquito host-seeking behavior.