Because of the avalability of a large amount of genetic information and rapidly accumulating molecular biology, the dense cluster of functionally-related genes in the DOPA deccarboxylase (Ddc) region (2-53.9+; 37Blo-D1) of the Drosophila melanogaster genome presents an unusual opportunity to examine the evolution and functional significane of tightly clustered genes that have evolved far enough from each other to specify separate, vital functins. Some of these genes code for catecholamine metabolizing enzymes required for neurotransmitter synthesis and sclerotization of the cuticle while others, also required for sclerotization and for female fertility, may specify key cuticle proteins. Most of the 18 genes located in the 160 kb of DNA in the Ddc region are located in two dense subclusters; the distal one with five genes in 15.5 kb includig Dox-A2 and the proximal one with nine genes in 25 kb including Ddc and amd. Several genes in the region share sequence homology indicating that many of the genes are evolutionarily related.- The analysis of the genetic and molecular organization of this cluster of vital, functionally- related genes will be completed. The evolutionary relationship and functioal significance of these genes will be investigated by searching for clustering of these genes in other Drosophila species and by mesuring mRNA levels of transposed genes to determine whether clustering is required for optimal expression. The structural and functional relatedness of these genes will be documented by sequencing a number of genes to determine the extent of sequence homology, by measuring stage and tissue specific mRNA levels and by determining how many of the genes respond to ecdysone. In order to characterize the specific function of individual genes, the effects of mutations on cuticle formation and female fertility in mutant: wild type chimeras will be examined, and catecholamine pools measured in mutant homozygotes. Information on specific biochemical function will be sought by comparing amino acid sequences with known sequences and by localizing protein sequences in cells using antibodies made against proteins proteins derived from expression vectors. ovary-specific control sequences will be sought using female sterile alleles along with transposed chimeric DNA constructs. Besides the evolutionary aspects, this research will make contributions to our knowledge of catecholamine metabolism, sclerotization, and cis-acting control elements.
