Proteomics of mosquito immune response to malaria

描述（由申请人提供）：疟疾媒介蚊子具有先天免疫系统，可以识别疟疾寄生虫感染并对其作出反应。血淋巴可能是蚊子最活跃的免疫区室，这是一种充满开放体腔的可溶性蛋白质血清。疟疾动合子、卵囊和子孢子必须存活，因为血淋巴中的免疫因子可以通过黑色素包囊或其他机制破坏它们。子孢子必须在血淋巴内迁移并侵入唾液腺才能传播。这项提案的母基金主要用于蚊子体内疟疾子孢子的生物学研究，特别是确定唾液腺入侵的受体。在母基金资助下的工作表明，暴露于血淋巴隔室会导致大多数子孢子通过未知机制迅速破坏，其中不到20%逃逸侵入唾液腺。然而，由于技术的限制，关键的血淋巴室一直是疟疾生命周期的一个很少研究的时期。血淋巴特别适合于蛋白质组学表征，因为这个关键区室中的功能性免疫应答是基于许多可溶性蛋白质（总共1，500 - 3，000种）之间的相互作用，这些蛋白质由各种细胞类型在其他地方合成，因此无法通过基于mRNA的测定在空间或时间上准确描述。拟议的项目将利用蛋白质组学的新技术进展以及寄生虫和载体的新基因组序列，从传染病载体中生成第一个（据我们所知）蛋白质组数据集。为此，我们提出了两个目标：1）利用液相色谱-串联质谱法对冈比亚按蚊血淋巴进行测序，鉴定静止状态下蚊子血淋巴的功能蛋白质组。2)进行差异表达分析以鉴定疟原虫子孢子感染诱导的血淋巴蛋白。生成的数据集将通过网络档案库发布并公开提供。了解蚊子的抗寄生虫免疫反应可以提供新的基于病媒的疟疾控制机会。此外，无脊椎动物的先天免疫反应显然是哺乳动物先天免疫的系统发育前体，越来越多的分子描述了结构和功能的保守性。这是可能的识别和信号转导机制参与先天免疫激活的子孢子在蚊子中可能是类似的保守在哺乳动物中，这可能是有助于疫苗的开发。

DESCRIPTION (provided by applicant): Malaria vector mosquitoes possess an innate immune system that can recognize and respond to infection by malaria parasites. Probably the most active immune compartment of the mosquito is the hemolymph, a serum of soluble proteins that fills the open body cavity. Malaria ookinetes, oocysts, and sporozoites must survive as yet uncharacterized immune factors in the hemolymph that can destroy them by melanotic encapsulation or other mechanisms. Sporozoites must migrate within hemolymph and invade the salivary glands in order for transmission to occur. The parent grant to this proposal focuses on the biology of malaria sporozoites in the mosquito, particularly the identification of receptors for salivary gland invasion. Work under the parent grant has shown that exposure to the hemolymph compartment causes rapid destruction of the majority of sporozoites by an unknown mechanism, which fewer than 20% escape to invade the salivary glands. Due to technological limitations, however, the critical hemolymph compartment has been a little-studied period of the malaria life cycle. Hemolymph is particularly well suited for proteomic characterization because the functional immune response in this key compartment is based on the interplay between many soluble proteins (1,500-3,000 total) that are synthesized elsewhere by a variety of cell types, and thus cannot be described accurately either spatially or temporally by mRNA-based assays. The proposed project will leverage new technological advances in proteomics and the new genome sequences of parasite and vector to generate the first (to our knowledge) proteomic datasets from an infectious disease vector. To this end, we propose two Aims: 1) Identify the functional proteome of mosquito hemolymph in the resting state using new approaches in liquid chromatography-tandem mass-spectrometry for sequencing of purified Anopheles gambiae hemolymph. 2) Carry out differential expression analysis to identify hemolymph proteins that are induced by malaria sporozoite infection. The generated datasets will be published and made publicly available by web archive. Understanding the mosquito anti-parasite immune response could offer new vector-based malaria control opportunities. Moreover, the innate immune response of invertebrates is clearly the phylogenetic precursor of mammalian innate immunity, with structural and functional conservation described for a growing number of molecules. It is possible that recognition and signaling mechanisms involved in innate immune activation by sporozoites in mosquitoes may be similarly conserved in mammals, which could be helpful in vaccine development.