Expression of the early-late gene encoding the nuclear receptor HR3 suggests its involvement in regulating the vitellogenic response to ecdysone in the adult mosquito

The insect steroid hormone, 20-hydroxyecdysone (20E), is a key factor contr olling critical developmental events of embryogenesis, larval molting, meta morphosis, and, in some insects, reproduction. We are interested in underst anding the molecular basis of the steroid hormone ecdysone action in insect egg development. The yellow fever mosquito, Aedes aegypti, in addition to being an important vector of human diseases, represents an outstanding mode l for studying molecular mechanisms underlying egg maturation due to string ently controlled, blood meal-activated reproductive events in this insect. To elucidate the genetic regulatory hierarchy controlling the reproductive ecdysone response, we have investigated ecdysone-regulated gene expression in vitellogenic mosquito ovaries and fat bodies. We have previously demonst rated the conservation of a primary ecdysone-triggered regulatory hierarchy , implicated in development of immature stages of Drosophila, represented b y the ecdysone receptor/Ultraspiracle complex and an early gene E75 during the reproductive ecdysone response (Wang, S.-F., Miura, K., Miksicek, R.J., Segraves, W.A., Raikhel, A.S., 1998. DNA binding and transactivation chara cteristics of the mosquito ecdysone receptor - Ultraspiracle complex. J. Bi ol. Chem. 273, 27531-27540; Pierceall, W.E., Li, C., Biran, A., Miura, K., Raikhel, A.S., Segraves, W.A., 1999. E75 expression in mosquito ovary and f at body suggests reiterative use of ecdysone-regulated hierarchies in devel opment and reproduction. Mel. Cell. Endocrinol. 150, 73-89). The present pa per demonstrates that conservation of the factors involved in the ecdysone- responsive genetic hierarchy regulating female reproduction extends beyond the early genes. Here, we identify AHR3, a highly conserved homologue of th e Drosophila HR3 early-late ecdysone-inducible gene in the mosquito. We sho w that AHR3 is expressed in both vitellogenic tissues of the female mosquit o, the fat body and the ovary. The expression of AHR3 correlates with the e cdysteroid titer, reaching a peak at 24 h after a blood meal. Moreover, in vitro fat body culture experiments demonstrate that the kinetics and dose r esponse of AHR3 to 20-hydroxyecdysone (20E), an active ecdysteroid in the m osquito, is similar to those of the late vitellogenic genes rather than the early E75 gene. However, as shown for other early and early-late genes, th e 20E activation of AHR3 is not inhibited by the presence of cycloheximide, a protein synthesis inhibitor. Taken together, these findings strongly sug gest AHR3 involvement in regulating the vitellogenic response to ecdysone i n the adult mosquito. (C) 2000 Elsevier Science Ireland Ltd. All rights res erved.