The proposed research is designed to advance our basic knowledge and understanding of the regulation of gene expression and enzyme synthesis in higher organisms. Two regulatory circuits of Neurospora, which control the expression of sulfur-related and nitrogen-related enzymes will be studied at the molecular level, with the use of genetically defined structural and regulatory genes. It is planned to identify the products of major regulatory genes which are postulated to encode DNA-binding proteins which are localized in the nucleus and have certain predicted characteristics. The binding of these regulatory proteins to DNA and to specific control recognition sequences will be studied, and the effect upon binding affinity of metabolic effectors will be determined. Using specific nitrogen and sulfur structural genes which will be cloned by a single efficient technique (in E coli) we plan to identify and sequence the control recognition sequences predicted to occur near the 5' end of each structural gene. The cloned genes will also be used as specific molecular probes to examine the importance of transcriptional control of the sulfur genes by the major sulfur regulatory gene and, similarly, of the nitrogen-related genes by the major nitrogen control gene.
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