De novo biosynthesis of uridine-5'-monophosphate (UMP) is performed by an unbranched pathway in animals that includes two multienzyme complexes and one mitochondrial enzyme. The basic structures and enzymatic functions of the pathway components appear to be essentially the same in all animals. Relatively little is known about genetic control of the pathway in animals; however, genes controlling all of the pathway enzymes have now been identified in Drosophila melanogaster. Understanding the organization and control of these genes in this animal will provide a model for genetic control of this pathway in all animals, including humans. An integrated group of studies is proposed to examine the genetic and molecular properties of the r, Dhod and r-1 genes and to seek functional and regulatory relationships among these loci. Noncomplementing r mutants will be mapped within this locus to localize regions coding for certain functions and r gene DNA will be cloned and sequenced to identify the molecular properties of mutant genes. Dhod null-activity mutants will be obtained and analyzed to determine the functions of the enzyme product of this gene. Mutants of the r-1 locus will be mapped within the locus and the protein product(s) of this locus will be purified and studied. Monoclonal antibodies against this protein(s) will be derived and used to isolate r-1 mRNA and cloned DNA. The latter will be analyzed and compared to accumulating information on cloned r DNA sequence organization. Genetic variants that affect multiple gene activities will be sought and studied. These experiments will be supported and complemented by parallel experiments into the expression of r, Dhod and r-1 genes in cultured Drosophila cells. Mutant cell lines will be obtained and together with extant r mutant cell lines, they will be used to study mutation reversion, DNA transformation and gene amplification phenomena.
