GENETIC REGULATION OF PYRIDINE DEHYDROGENASES

6-磷酸葡萄糖脱氢酶 脱氢酶、异柠檬酸脱氢酶和苹果酸酶份额 一种常见的作为辅酶的吡啶核苷酸(NADP)。的活动 这些酶被协调起来，以产生足够的 NADPH用于脂质生物合成。在生物化学方面的比较 吡啶连接脱氢酶的参数表明一个共同的 这些酶的进化起源。 在黑腹果蝇中，似乎有两种形式的 NADP-异柠檬酸脱氢酶和可能有两个基因 控制这种酶的活性。一种分子 该基因的特征(S)及其转录产物(S) 被提议作为理解这一规则的第一步 基因-酶系统。果蝇基因表达文库 将对IDH特异序列进行筛选。的身份 这些克隆的序列将通过原位杂交进行验证 唾液腺聚乙烯染色体及其免疫沉淀法 体外翻译的杂交种筛选出的mRNA。克隆是 由这些技术识别的将被用于分离 相应的基因组DNA序列。制约分析 将比较cdna和基因组序列。发展中的 这些序列的表达将由Southern决定 分析。TO基因链特异性转录本的杂交 适当发育的蛹mRNAs的“Northern”印迹将允许 转录方向的确定。未来 实验将包括S1核酸酶图谱和引物 可拓分析。

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.