An understanding of mechanisms regulating gene expression in neurons is central to the understanding of neuronal development and function and to the analysis of disease processes as they affect neurons. Of particular interest are those genes which are unique to neurons or whose expression distinguishes neurons from other kinds of cells. The long-term objective of this research program is to elucidate through the techniques of molecular genetics those basic features of the DNA sequence and organization of neuronal genes which regulate and limit their expression to neurons. As a first step, the mechanisms controlling the expression of genes for the neurofilament will be investigated. Three genes specify proteins which comprise neurofilaments. These genes are active at high levels in neurons and are completely inactive in all other cell types in the body. The activities of these genes are discoordinately controlled during embryonic development but change coordinately when axons are damaged. The molecular mechanisms responsible for the control of these genes are expected to be complex and to involve the interaction of protein factors with specific regulatory sequences within the DNA of the genes. This project will use recombinant DNA clones and DNA sequence information already available to identify those regulatory sequences, and to search for postulated protein regulatory factors which bind to the DNA. To do this we propose to locate the major block of enhancer elements of the NF (M) and NF (H) genes and establish their 3' and 5' boundaries. We will demonstrate that these sequences are sufficient to specify neuron specific gene expression by investigating the expression in transgenic mice of hybrid genes containing these enhancer elements linked to simple reporter genes. Next we propose to identify component sequence motifs within the enhancer that are important for expression during differentiation of embryonal carcinoma cells to neuron-like cells or for neuron specific expression in transgenic mice. Ultimately, we will characterize the proteins that bind to these component motifs and obtain cDNA clones of them. Accumulations of neurofilaments or neurofilament-like proteins occur in a number of human disorders of the nervous system, including Alzheimer's and Parkinson's disease. The information obtained during this study on neurofilament gene regulation will constitute fundamental information for analyzing the molecular basis of pathological neurofilamentous changes in human.
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