Glucose 6-phosphate dehydrogenase, 6-phosphogluconate dehydrogenase, isocitrate dehydrogenase and malic enzyme share a common pyridine nucleotide (NADP) as coenzyme. Activities of these enzymes are coordinated to generate adequate pools of NADPH for lipid biosyntheses. Biochemical comparisons of the parameters of pyridine-linked dehydrogenases suggest a common evolutionary origin of these enzymes. In Drosophila melangaster there appears to be two forms of NADP-isocitrate dehyrogenase and there may be two genes controlling the activity of this enzyme. A molecular characterization of this gene(s) and the transcript(s) made from it is proposed as the initial step in understanding regulation of this gene-enzyme system. Expression libraries of Drosophila cDNA will be screened for sequences specific for IDH. The identity of these cloned sequences will be verified by in situ hybridization to salivary gland polythene chromosomes and immunoprecipitation of in vitro translated hybrid selected mRNA. Clones which are identified by these techniques will be used to isolate corresponding sequences of genomic DNA. Restriction analyses will compare cDNA and genomic sequences. The developmental expression of these sequences will be determined by Southern analysis. Hybridization of strand specific transcripts of cDNA to """"""""Northern"""""""" blots of appropriately staged pupal mRNA will allow determiniation of the orientation of transcription. Future experiments would include S1 nuclease mapping and primer extension analysis.
