Analysis of the Inverse Effect in Drosophila

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The long range goal of the proposed work is to understand the nature and mechanism of regulatory genes that inversely affect the expression of the white eye color gene. It is believed that this type of regulatory effect is involved in the expression of most types of structural genes. The four currently known inverse regulators of white will be precisely localized, genetically and cytologically, via x-ray and P element mutagenesis as a prelude to molecular cloning. Cloning of two of these genes will reveal their nature and provide the means to address the molecular mechanism of action on white by sequence analysis, in vitro binding tests and genetic dosage studies with transformed copies. Each of the four inverse regulators, individually as well as in combinations, will be tested for an effect upon white mRNA levels in developmental profiles, as well as for a coordinate response from scarlet and brown, which cooperate with white in pigment deposition. Each will be tested for an effect on a white promoter-reporter construct to examine the requirement of the w promoter. A refined functional analysis of the white promoter will be conducted to better localize the sites required for interaction with the four regulators. This analysis will be conducted by deletion and site directed mutagenesis of the promoter, which will then be placed driving the alcohol dehydrogenase structural gene. These constructs will be tested to determine the four inverse regulators and examine the context within which these regions work. The proposed experiments will contribute to an understanding of the inverse regulation phenomenon and address whether there is a connection between such regulatory genes and the biological phenomena associated with chromosomal dosage such as compensation, the inverse effect, aneuploid syndromes, and position effect variegation.*** //