Hemocyte Transcriptome and Immunity in Aedes aegypti

蚊子的先天免疫反应是控制病媒与病原体相互作用的主要因素； 因此，它是媒介能力的主要决定因素。蚊子和其他昆虫的先天免疫 使用细胞和体液成分来应对入侵的病原体，但对体液方面的研究 天然免疫的主要成分，主要侧重于抗菌肽(AMP)和 参与其产生的信号通路。对血细胞和细胞的重视要少得多。 免疫反应，但最近的数据清楚地证实，细胞反应可能在清除微生物方面发挥关键作用 病原体通过吞噬作用、黑化作用或两者的结合作用而致病。所需的数据中存在一个空白 准确定义调节过程、血细胞产生的效应分子以及这些细胞和细胞的作用 产品在先天免疫力中发挥作用。同样清楚的是，AMP是针对以下疾病的先天免疫反应的重要因素 入侵的微生物，但尚不清楚细胞和基于AMP的体液反应如何协同作用来清除 微生物感染。我们认为，一种简化论的方法，研究一个基因，或一小组基因，与 对于一种表型，在确定蚊子先天存在的复杂相互关系方面存在局限性 免疫反应。因此，为了开创更全面的研究先天免疫的实验方法。 蚊子，我们开发了埃及伊蚊免疫激活的大型表达序列标签(EST)数据集 血细胞和基于互补寡核苷酸的微阵列作为底物也用于评估蚊子细胞 作为体液免疫反应。这里提出的研究的一般假设是蚊子天生 免疫反应是一个高度复杂的过程网络，它使用截然不同但不相互排斥的 吞噬和黑化结合AMPS对清除猪血淋巴中细菌感染的反应 被感染的蚊子。为了验证这一假设，我们将使用蚊子Ae。埃及伊蚊和一些细菌种类 (1)高通量分子方法，以描述基因转录并识别潜在的不同模式 吞噬和黑化的抗细菌反应，(2)利用微阵列分析来评估转录图谱 与不同细菌种类(其致病性和引起反应的能力不同)有关的变化 为了更好地剖析影响细胞免疫的机制，以及(3)使用微阵列分析，合成肽， RNAi方法和表型评估，以更好地定义AMP的作用，并选择未知基因，在 先天免疫力。所有来自微阵列分析的数据，包括使用的实验方案，都将被制作 在包含我们的血细胞EST数据的互动网站上公开提供。

The mosquito innate immune response is a major factor governing the interaction between vector and pathogen; consequently, it is a primary determinant of vector competence. Innate immunity in mosquitoes and other insects employs both cellular and humoral components in response to invading pathogens, but studies of the humoral aspects of innate immunity have dominated, with a primary emphasis on the identification of antimicrobial peptides (AMPs) and the signaling pathways involved in their production. Far less emphasis has been placed on hemocytes and cellular immune responses, but recent data clearly verify that cellular responses likely play a critical role in clearing microbial pathogens by phagocytosis, melanization, or by both processes in combination. There is a void in the data required to accurately define regulatory processes, hemocyte-produced effector molecules, and the role these cells and cell products play in innate immunity. It also is clear that AMPs are significant factors in innate immune responses against invading microbes, but it is not known how cellular and AMP-based humoral responses function in concert to clear microbial infections. We believe that a reductionist approach, that studies one gene, or a small set of genes, in relation to a phenotype, has limitations for determining the complex interrelationships that undoubtedly exist in mosquito innate immune responses. Therefore, in order to initate more holistic experimental approaches for studying innate immunity in mosquitoes, we developed a large expressed sequence tag (EST) data set from Aedes aegypti immune-activated hemocytes and complementary oligonucleotide-based microarrays as substrates to evaluate mosquito cellular, as well as humoral, immune responses. The general hypothesis for the research proposed herein is that mosquito innate immune responses are a highly complex network of processes that employ the distinct, but not mutually exclusive, responses of phagocytosis and melanization, in concert with AMPs, to clear bacteria infections from the hemolymph of infected mosquitoes. To test this hypothesis, we will use the mosquito Ae. aegypti and a number of bacteria species in (1) high-throughput molecular approaches to profile gene transcription and to identify distinct patterns underlying the anti-bacterial responses of phagocytosis and melanization, (2) utilize microarray analyses to assess transcription profile changes in relation to different bacterial species (that vary in their pathogenicity and ability to elicit a response) in order to better dissect mechanisms influencing cellular immunity, and (3) employ microarray analyses, synthetic peptides, RNAi methodologies, and phenotype evaluations to better define the role AMPs, and select unknown genes, play in innate immunity. All data resulting from microarray analyses, including experimental protocols used, will be made publicly available on the interactive website that contains our hemocyte EST data.