Mammals employ a complex series of recombination events (V(D)J recombination) to assemble the enormous diversity of antigen receptors required to mount an effective immune response. Proper targeting of these events is pivotal to lymphopoiesis; aberrations can produce chromosomal translocations that result in lymphoid tumors. A single V(D)J recombinase activity mediates all recombination events by recognizing sequences common to all antigen receptor gene segments. Despite these shared features, receptor gene assembly is tightly regulated at the levels of tissue-, stage- and allele-specificity. Current evidence suggests that specificity is imposed by modulations in accessibility of chromosomal gene segments to recombinase. The molecular determinants of recombinational accessibility; however, have not been delineated. The objective of this proposal is to elucidate the mechanisms by which recombinase activity is properly targeted to distinct genetic loci during lymphocyte development. Specifically, the proposed research will: l) Independently assess the roles of transcription and enhancer activity in providing V(D)J recombinational accessibility; 2) Elucidate the role of NFkappaB in assembly and expression of antigen receptor loci; 3) Determine the minimal requirements for activation of a recombinationally inert receptor locus; 4) Define genetic elements that confer tissue-specific targeting of recombinase activity. Collectively, these studies will yield fundamental insights into the molecular elements that initially direct recombinase activity to particular antigen receptor loci, and subsequently re-direct this activity in order to ensure lymphocyte mono-specificity. Such insights are expected to provide a greater understanding of mechanisms that confer differential, long-range regulation to complex genetic loci. Dr. Oltz has studied the mechanisms and regulation of V(D)J recombination extensively during his training as a molecular immunologist. The initial goals of this research proposal will provide a view of recombinational accessibility at an intermediate level of molecular resolution. Dr. Oltz will expand these findings in two complementary directions. At an increased molecular level, he will pursue the precise features of transcription, enhancer activity and chromatin structure that contribute to recombinational accessibility. Conversely, at a physiological level, animal models will be used to examine the significance of these observations in normal developing lymphocytes. The interactive environment, and diverse expertise offered by Dr. Oltz's colleagues at Vanderbilt, and its Cancer Center, provide an ideal scientific setting to foster these broadened approaches.
