It is the long term objective of these studies to learn about the genomic structures required for the expression of particular proteins in particular neuronal and glial cells and the factors which regulate that expression. The hypothesis generally accepted by molecular biologists that trans-acting factors act at specific sequences to stimulate transcriptional initiation. The factors and the sequences at which they act are known for only a few genes at present. Many proteins of the central nervous system (CNS) are thought to be expressed in only a subset of neurons. Dr. Sutcliffe specific hypothesis will test the particular sequences or sets of sequences determine which genes are actively transcribed in particular cells in response to the set of trans-acting transcriptional stimulatory factors present in those cells. The specific studies described in this proposal will involve characterization at the nucleotide sequence level of the 5' ends of the mRNAs and the regions upstream from the transcriptional initiation sites on the genes for neuron- specific enolase and non-neuronal enolase. These studies will define the promoter regions for these two genes so that appropriate transfection assays for their function can be designed. Those assays will allow deletion and mutagenesis experiments to be performed which will indicate the particular nucleotides required for specifying brain cell-specific gene expression, and will provide information necessary for identifying the trans-acting factors which regulate transcription of these two genes. Both promoter and enhancer elements will be investigated. These studies should provide insights into the specific mechanisms which regulate transcription of these particular genes and also illuminate the features of general classes of gene regulatory sequences which determine developmental stage- and lineage-specific gene expression in the brain.
