The molecular and cellular basis of circadian rhythms in vector mosquitoes

PROJECT SUMMARY/ABSTRACT Mosquito disease transmission relies on the insect's ability to feed on humans. Like other animals, vector mosquitoes exhibit circadian patterns of behaviors, including blood feeding. Although mosquitoes are known to possess a canonical molecular clock that controls their patterns of activity and blood feeding, little is known about the cells in which clock genes act or the manner in which mosquito clocks operate at a molecular level, given their apparent differences from the well-studied mammalian and fly clocks. This proposal will identify and molecularly characterize the molecular clock and clock neurons of two vectors of human disease, the nocturnal malaria mosquito Anopheles gambiae (An. gambiae) and the crepuscular dengue mosquito Aedes aegypti (Ae. aegypti). It will test how disruptions in core clock genes alter the molecular properties and functions of circadian neurons and impact behavior. It will also explore the roles of clock genes in peripheral tissues, focusing on sensory transduction. Aim 1) Identify cells expressing core clock genes in Ae. aegypti and An. gambiae: Genetic access will be obtained to cells expressing core clock genes by creating QF2 knock-ins into key clock genes, including per, Clk, tim, M-cry and D-cry. These will permit assessment of how each contributes to the function of the mosquito clock and whether they act in overlapping or distinct cell populations. Aim 2) Define the cellular composition of mosquito clocks and the impact of clock genes on their gene expression patterns: scRNA-seq based analyses will be performed across the circadian cycle to create a comprehensive atlas of the circadian neurons of the Ae. aegypti and An. gambiae brains, and examine how the loss of core circadian genes impacts these patterns of gene expression. Aim 3) Determine the contribution of the circadian clock to sensory detection and behavior: Loss- of-function mutations in core clock genes will be used to assess how their activity affects circadian patterns of locomotion, light responses and blood feeding, as well as the detection of host-associated sensory cues.