This proposal is to identify and purify the inhibitor protein (PKI) of the cAMP-dependent protein kinase A in fruit flies. A purification scheme similar to that used for mammalian PKI will be employed. Fruit flies have a relatively simple genome and many mutants have been described, making them useful experimental organisms. After the PKI is purified, primers will be prepared in order to obtain and sequence the gene. Gene expression in various tissues and at various stages of development will be determined. The PKI gene will be mutated to determine the resulting phenotype. Potential roles of this gene in embryonic development, especially in the nervous system and in pattern formation, will be examined. Previously described mutants have been shown to have abnormal levels of cAMP. The proposed experiments will help elucidate the role of PKI in regulating cAMP-dependent processes in fruit flies. %%% Mutants of fruit flies have been described which have altered cAMP levels. cAMP-dependent protein kinase (PKA) activity is involved in many cellular processes, changing the behavior of target proteins. This proposal is to isolate and study the properties of a protein (PKI) which inhibits PKA activity. The gene for PKI will then be isolated, mutations will be made, and the phenotype of the flies will be determined. Gene expression in various tissues and at various times in development will be measured. The information obtained should increase understanding of what roles PKA and PKI play in development and pattern formation, and why regulation of cAMP levels seems to be critical for many processes in fruit flies.
