SERINE PROTEASES AND IMMUNITY IN ANOPHELES GAMBIAE

疟疾是一种由疟原虫属原生动物引起的疾病，通过蚊子传播给人类。每年，数以亿计的人感染这种微生物，导致200-300万人死亡，其中大部分是幼儿。这一问题在非洲最为严重，由于寄生虫对过去用于预防和控制的廉价药物的抗药性日益增强，该问题现在正在恶化。另一项重要的控制战略--通过使用杀虫剂减少蚊子数量--也因杀虫剂抗药性、成本增加和训练有素的人员流失而受到威胁。一个新的研究目标是蚊子和寄生虫之间的相互作用。这项研究的长期目标是了解蚊子和疟疾寄生虫之间不相容的分子基础，因为这可能会提出一些方法，通过对媒介蚊子的遗传或化学操作来增强导致不相容的机制。这项拟议的研究将集中在鉴定参与蚊子体液免疫反应的蛋白质和基因，体液免疫反应可能导致寄生虫死亡。特别是，丝氨酸蛋白酶是产生两种杀死寄生虫的机制的重要酶，即黑色素包裹和抗菌反应。在非洲疟疾媒介冈比亚按蚊中，三个丝氨酸蛋白酶基因AgSp14D1、AgSp14D2和AgSp14A在注射细菌或感染疟疾寄生虫后显示转录水平的变化。这些分子将在体外和体内进行表征和测试，以参与针对寄生虫的免疫反应。此外，在触发寄生虫黑化过程中起重要作用的酚氧化酶原激活丝氨酸蛋白酶将被生化提纯，然后克隆和鉴定。最后，几种冈比亚按蚊丝氨酸蛋白酶的结构类似于其他参与免疫反应的酶，将在mRNA和蛋白质水平上进行表征。

Malaria, a disease caused by parasitic protozoans in the genus Plasmodium, is transmitted by mosquitoes to humans. Each year, hundreds of millions of people are infected with this organism and 2-3 million fatalities result, mostly in young children. The problem is most severe in Africa, and is now worsening because of the increasing resistance of parasites to the inexpensive drugs used for prevention and control in the past. A second important strategy for control, reducing mosquito populations through insecticide use, is also threatened because of insecticide resistance, increasing costs and loss of trained personnel. A new target for research has been the interaction between the mosquito and the parasite. The long-term goal of the research presented here is to understand the molecular basis for incompatibility between mosquitoes and malaria parasites because this may suggest ways to enhance mechanisms that cause incompatibility through genetic or chemical manipulation of vector mosquitoes. The proposed research will focus on characterizing proteins and genes that are involved in the mosquito's humoral immune responses, which can lead to the death of parasites. In particular, serine proteases are important enzymes in generation of two parasite-killing mechanisms, melanotic encapsulation and the antimicrobial response. In the African malaria vector, Anopheles gambiae, three serine protease genes, AgSp14D1, AgSp14D2, and AgSp14A, show changes in transcript levels after bacterial injections or infection with malaria parasites. These molecules will be characterized and tested for involvement in immune responses against parasites in vitro and in vivo. In addition, prophenoloxidase activating serine proteases, which are important in triggering parasite melanization, will be biochemically purified and then cloned and characterized. Finally, several A. gambiae serine proteases that exhibit domain structures similar to other enzymes involved in immune responses will be characterized at the mRNA and protein levels.