Glucocorticoids regulate the expression of a number of genes in mammals. An interesting aspect of this regulation is that hormonal stimulation can induce the expression of different pools of genes in different tissues. The goal of the proposed study is to obtain a better understanding of the general mechanisms of expression and inducibility of glucocorticoid regulated genes in mammals and, more specifically, to determine the role of chromatin structure in determining the inducibility of tissue-specific gene pools. Two glucocorticoid regulated genes in the rat will be used. The genes encoding tyrosine aminotransferase (rTAT-synthesized in the liver) and growth hormone (rGH synthesized in pituitary) will be studied in cultured HTC (rat hepatoma) and GC (rat pituitary tumor) cells. The general chromatin properties of these genes (nuclease sensitivities, presence of ordered nucleosomal arrays and nuclear matrix attachment) will be contrasted for cells in which the genes are expressed and glucocorticoid inducible to cells in which they are not in order to identify different chromatin domains associated with expression and hormone action. The span of domains identified in this way will be determined by chromosomal walking. Using DNA-mediated gene transfer, the DNA sequences located at the junctions of the rTAT and rGH domains will be tested for their ability to direct or delimit the establishment of domains of altered chromatin structure. In addition a limited analysis of nucleosome composition and DNA methylation of sequences spanning these junctions will be made. The long term goal of this research is to characterize factors responsible for the assembly and/or maintenance of chromatin structural domains in mammals. These findings should be helpful for describing not only the molecular biology of glucocorticoid induction of gene expression, but also more general processes of cellular development and differentiation.
