A universally conserved property of long-term memory (LTM) formation, is the requirement for a cascade of gene expression under control of the cAMP responsive enhancer binding protein (CREB). However in no case have CREB activated LTM genes been identified in a system which is amenable to genetic analysis. Thr primary goal of the research proposed here is to isolate and characterize CREB target genes in Drosophila, and begin an analysis of their role in LTM. The rich array of molecular and genetic approaches available in Drosophila, as well s the availability of established standard assays for long-term associative memory, make Drosophila an attractive system for this project. CREB target genes will be identified using a technique known as differential mRNA display. This is a PCR based technique which as been successfully used to isolate mRNAs that are expressed differently in two tissues of interest. In this case, differential display will be applied to mRNA isolated from brain tissue from two different groups of flies. In the first group, a transgene encoding a CREB blocker isoform will be induced, a procedure which as been shown to specifically block LTM (see appendix). In the second group, a transgene encoding a CREB activator isoform will be induced. This second procedure has been shown to enhance LTM formation. PCR fragments isolated in this screen will be used for RNAase protection and insitu hybridization analyses to confirm CREB/LTM responsive regulation. Candidate fragments will then be used to screen brain CDNA libraries using standard techniques. The role of novel cDNAs in LTM formation will then be tested by standard miss-expression and mutagenesis approaches.
