These studies will extend our understanding of mosquito reproduction and provide a basis for novel genetic and physiological strategies for mosquito control. The long term goals are 1) to identify and understand at the molecular level, key physiological processes that control mosquito reproduction, 2) to clone genes whose expression is essential to those processes, and 3) to use the cloned genes to develop effective gene transfer procedures. The genetic approaches that will be extended to the mosquito have an immediate application for asking detailed questions about gene expression, and long term application to the genetic control of mosquito populations. The importance of the reproductive cycle in disease maintenance and transmission, and the fact that the mosquito Aedes aegypti has served as a primary model for understanding reproduction in insects, contributed to the choice of this system. Our research is focussed on the cycle of ribosome synthesis and degradation that accompanies vitellogenin synthesis. In Drosophila, genetic disruption of ribosome biosynthesis has been shown to interfere with egg production, indicating that the ribosome is a potential target for genetic manipulation of mosquito reproduction.
Our specific aims i nclude the use of molecular and physiological approaches to describe ribosome biosynthesis within the context of the endocrine regulation of mosquito reproduction. Cloned rRNA and ribosomal protein genes are available for use as probes in this study. The cloned genes will be characterized, with specific emphasis on identifying promoter sequences. This work will include development of an in vitro transcription system, which provides a tool for eventual identification of specific factors that modulate rRNA gene expression. Promoters will also be defined using functional assays in transfected cells. The promoters from ribosomal protein genes will be used to engineer DNA plasmids with specificity for expression in mosquito cells. A somatic transformation assay will be used to express, in the intact mosquito, DNA constructs whose expression was first optimized in cultured cells. A selection system that will facilitate rapid genetic screening of transformed mosquitoes will be developed. These studies will contribute to an increased understanding of the genetics and physiology of the mosquito, a disease vector with considerable impact on human health and productivity.
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