The role of circadian clock genes in the overwintering diapause of the northern house mosquito, Culex pipiens

Most insects in temperate environments use photoperiod as a token stimulus for the advent of winter, and enter diapause. However, the molecular mechanisms by which insects interpret day length are not yet understood. Researchers have long suspected that circadian clock genes, which provide insects with information on light:dark cycles, might also be involved in the diapause response. The Northern House Mosquito, Culex pipiens, is the major vector of West Nile Virus. During winter, adult female mosquitoes enter a reproductive diapause which is characterized by arrested egg follicle development. To determine whether canonical circadian genes, such as period, timeless, Cycle and Clock play a role in diapause initiation in Culex pipiens, we measured their expression pattern over a 24-hour period in adult females that were reared under diapausing conditions (short day 18C), or non-diapausing conditions (long day at 25C or 18C). Gene expression in diapausing C. pipiens was measured 1 week and 1 month after eclosion, while non-diapausing mosquitoes were sampled one week after adult emergence. To further implicate the involvement of clock genes in the diapause response, we knocked down the expression of period and Cycle using RNA interference. Both diapausing and non-diapausing female mosquitoes were injected with either period, Cycle or B-gal dsRNA or insect saline on the day of eclosion; egg follicle length was measured 14 days later. Our results suggest that period is involved in diapause initiation as period-injected females that were reared under diapausing conditions had egg follicles that were significantly larger than insect saline-injected controls.