Molecular regulation of diapause in Culex pipiens

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DESCRIPTION (provided by applicant): Mosquitoes, like most insects, are able to feed and reproduce only a few months each year. The remaining months are spent in a form of dormancy known as diapause. It is the objective of this proposal to understand the molecular mechanisms that regulate this developmental arrest in adults of the northern house mosquito, Culex pipiens, the vector of West Nile virus, with the longterm goal of disrupting this vulnerable phase of the life cycle. The three specific goals of this proposal focus on the molecular pathway that programs the diapauses state. Insulin signaling is a key intermediary in generating the diapause phenotype (traits such as a halt in development, fat accumulation, extended lifespan and increased stress resistance), and this project investigates pathways both up- and downstream of insulin signaling. Our first goal is to identify the pathways leading to the diapause phenotype by working downstream of insulin and FOXO (forkhead transcription factor). Using a FOXO antibody and ChIP sequencing, genes downstream of FOXO will be identified and their putative roles in diapause will be evaluated with functional assays including RNA interference (RNAi). The second goal, upstream of insulin signaling, is to identify the role of clock genes in the transduction of the short day length signals that are used by the mosquito to program entry into diapause. The major clock genes from C. pipiens will be cloned, expression patterns will be evaluated under both long day lengths (nondiapause-inducing) and short day lengths (diapause-inducing), and their function in programming diapause will be evaluated using RNAi. The third aim is to link the photoperiodic response to the downstream insulin/FOXO pathway. After disrupting the clock mechanism we will monitor the impact of these changes on gene expression patterns in the insulin/FOXO pathway. With this information, it will be possible to begin construction of the pathway leading from perception of photoperiod to execution of the diapause phenotype. Any alterations in the diapause response (e.g. diapause prevention, delay in onset or termination, impairment of energy storage or utilization, reduced stress tolerance) could be used as tools for disrupting this critical phase of the mosquito life cycle.