The function(s) of serpin-2 in mosquito immunity and physiology

Serpin-2 在蚊子免疫和生理学中的功能

• 批准号: 8163820
• 负责人: Kristin Michel
• 金额: $ 38.18万
• 依托单位: KANSAS STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-05-01 至 2015-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8163820
• 关键词: Adult; Advanced Development; Affect; African; Aftercare; Anopheles gambiae; Arthropods; Binding; Biochemical; Biochemical Genetics; Biochemistry; Biological; Cessation of life; Cleaved cell; Clinical; Clip; Complex; Crystallography; Culicidae; Data; Development; Double-Stranded RNA; Emerging Communicable Diseases; Female; Generations; Goals; Health; Hemolymph; Human; Immune; Immune response; Immune system; Immunity; Injection of therapeutic agent; Insecta; Insecticide Resistance; Insecticides; Knowledge; Laboratories; Lead; Life Tables; Longevity; Malaria; Measures; Methodology; Microfluidics; Mission; Modeling; Molecular; Molecular Target; Natural Immunity; Nature; Parasites; Peptide Hydrolases; Pharmaceutical Preparations; Phenotype; Physiological; Physiology; Population; Populations at Risk; Prevention; Proteins; Proxy; Public Health; Research; Resistance; Resistance development; Serine Protease; Serine Proteinase Inhibitors; Serpins; Site; Structure; Techniques; Testing; Time; United States National Institutes of Health; Vector-transmitted infectious disease; base; design; disease transmission; disorder control; extracellular; in vivo; inhibitor/antagonist; innovation; killings; mathematical model; mortality; novel; pathogen; pressure; prevent; pro-phenoloxidase; programs; reproductive; resistance mechanism; serpin-2; small molecule; success; transmission process; tumor; vector; vector control

DESCRIPTION (provided by applicant): Malaria control and eradication strategies continue to primarily rely on vector control that is stymied by insecticide resistance in vector populations. In addition to resistance management, novel insecticides with new modes of action - particularly for which efficacy is not affected by existing resistance mechanisms - are required. The long-term goal is to develop a malaria vector control methodology based on a new insecticide target, the serine protease inhibitor serpin (SRPN)2 from the African malaria mosquito, Anopheles gambiae s.s.. SRPN2 is a key negative regulator in the extracellular proteinase cascade that controls activation of prophenoloxidase (proPO) and thus melanization - a powerful, arthropod-specific innate immune response. SRPN2 depletion from the hemolymph of adult female mosquitoes significantly reduces longevity with escalating daily mortality nine days after treatment. The objectives in this application are to identify the interactions of SRPN2 with its proteinase targets and to evaluate the malaria control potential of a SRPN2 inhibitor. The rationale for the proposed research is that detailed information on SRPN2's biological proteinase targets and mode of action will ultimately allow the design of small molecule inhibitors of SRPN2 that could act as an insecticide for mosquito vector control. Guided by our preliminary data, the following three specific aims will be pursued: (1) Determine the molecular targets of SRPN2 inhibitory function; (2) Determine the molecular interface between SRPN2 and its target proteinase(s); and (3) Model the impact of SRPN2 depletion on disease transmission. Under the first aim, a combination of biochemical and genetic approaches that have been used successfully in the applicant's laboratory will be used to identify serine proteinase targets of SRPN2 based on their ability to (a) form covalent complexes with the serpin and (b) revert the serpin's depletion phenotype. Under the second aim, the molecular interactions between SRPN2 and one of its target proteinases, CLIPB9, will be identified by protein crystallography and mutational analysis. Under the third aim, the potential effect of SRPN2 inhibition on malaria transmission and resistance development against a potential SRPN2 inhibitor will be assessed by mathematical modeling. The proposed research is innovative, as it will for the first time evaluate the mosquito immune system as a physiological target for novel insecticides. Additionally, this project will use microfluidics as a highly innovative approach to the study of mosquito innate immunity, which if successful will be transformative to the field of insect biochemistry. This project is significant as it will provide fundamental knowledge of the nature of serine proteinase cascades that regulate melanization in mosquitoes. Ultimately, it has the potential to advance the development of new a generation of insecticide targets for malaria control. PUBLIC HEALTH RELEVANCE: Vector-borne diseases, especially malaria, continue to be a major public health threat world-wide, with roughly half of the world population at risk of malaria. The proposed project is relevant to public health because the identification of novel insecticide targets in mosquitoes will allow the development of new vector control measures for malaria control. Therefore, the proposed project is relevant to the NIH-NIAID's mission in relation to the understanding and prevention of re-emerging infectious diseases.

描述（由申请方提供）：疟疾控制和根除战略仍然主要依赖于病媒控制，但病媒控制因病媒种群对杀虫剂的抗药性而受阻。除了抗药性管理之外，还需要具有新作用模式的新型杀虫剂，特别是其功效不受现有抗药性机制影响的杀虫剂。长期目标是开发一种疟疾病媒控制方法，其基础是一种新的杀虫剂靶标，即来自非洲疟疾蚊子冈比亚按蚊的丝氨酸蛋白酶抑制剂serpin（SRPN）2。SRPN 2是细胞外蛋白酶级联中的关键负调节因子，其控制酚氧化酶原（proPO）的活化并因此控制黑化-一种强大的节肢动物特异性先天免疫应答。SRPN 2从成年雌性蚊子的血淋巴中消耗显著降低寿命，在处理后9天每日死亡率上升。本申请的目的是鉴定SRPN 2与其蛋白酶靶标的相互作用，并评估SRPN 2抑制剂的疟疾控制潜力。拟议研究的基本原理是，关于SRPN 2的生物蛋白酶靶标和作用模式的详细信息最终将允许设计SRPN 2的小分子抑制剂，其可以作为蚊子病媒控制的杀虫剂。根据我们的初步数据，将追求以下三个具体目标：（1）确定SRPN 2抑制功能的分子靶点;（2）确定SRPN 2与其靶蛋白酶之间的分子界面;（3）模拟SRPN 2缺失对疾病传播的影响。在第一个目标下，已经在申请人的实验室中成功使用的生物化学和遗传方法的组合将用于基于它们（a）与丝氨酸蛋白酶抑制剂形成共价复合物和（B）恢复丝氨酸蛋白酶抑制剂的耗尽表型的能力来鉴定SRPN 2的丝氨酸蛋白酶靶标。在第二个目标下，SRPN 2和其靶蛋白酶之一CLIPB 9之间的分子相互作用将通过蛋白质晶体学和突变分析来鉴定。在第三个目标下，将通过数学建模评估SRPN 2抑制对疟疾传播和对潜在SRPN 2抑制剂的抗性发展的潜在影响。这项研究具有创新性，因为它将首次评估蚊子免疫系统作为新型杀虫剂的生理靶点。此外，该项目将使用微流体技术作为研究蚊子先天免疫的高度创新方法，如果成功，将对昆虫生物化学领域产生变革。这个项目是重要的，因为它将提供丝氨酸蛋白酶级联调节蚊子黑化的性质的基础知识。最终，它有可能推动新一代疟疾控制杀虫剂目标的开发。 公共卫生关系：病媒传播疾病，特别是疟疾，仍然是世界范围内的一个主要公共卫生威胁，世界人口约有一半面临疟疾风险。拟议的项目与公共卫生有关，因为确定蚊子中的新杀虫剂目标将有助于制定新的疟疾病媒控制措施。因此，建议的项目是相关的NIH-NIAID的使命，在了解和预防重新出现的传染病。