The B-cell developmental system is a paradigm for understanding the mechanisms that enable an organism to respond to extracellular signals. Resting B-cells respond to antigenic stimuli by secretion of immunoglobulin molecules. One of the hallmarks of this terminal differentiation events is an increase in the expression of immunoglobulin molecules reflected in the increased levels of mRNA. Although a large number of studies have been exerted in elucidating the molecular basis for this alteration in the level of gene expression, we still know very little about the mechanisms involved in this process. Our studies of the mouse kappa immunoglobulin locus has revealed that the structure of chromatin around the 3' enchancer region is remarkably different in antibody-secreting plasma B-cells compared with that of mature B-cells. The DNAse I hypersensitive structure first observed in pro B-cells persists at the mature B-cell stage but undergoes a transformation at some stage between the differentiation of B-cells to plasma cells resulting in the spreading of nuclease susceptible sites upstream and downstream of the initial site. Since similar changes are detected in terminally differentiated plasma cells expressing lamba chains, these results suggest that the alteration in the 3'region is a differentiation- dependent event. These changes in chromatin structure may underlie an important aspect of B-cell end-differentiation an might bear upon kappa gene expression. By a combination of reverse genetic, immunological, cell biological, molecular biological, and biochemical techniques, we hope to identify the molecular basis of these chromatin structural changes. We have the unique opportunity to identify the factors that participate in terminal differentiative events in B-cell development, the external agents that these factors respond to, and their possible mechanism of action on gene expression. Specifically, we wish to address the following specific aims: 1) To determine the effects of extracellular stimuli (mitogens and cytokines) on the chromosomal alterations observed during B-cell maturation. This should lead to the establishment of an in vitro system for studying the mechanisms involved in 3' enhancer induction. 2) To localize the DNA sequences within the 3" enhancer that define the formation of DNAse I hypersensitive sites. 3) To identify the precise nucleotide sequences in contact with protein factors in vivo genomic footprinting. 4) To identify and biochemically characterize the protein factors that bind the DNA sequences identified in specific aim#2.