The specialized recombination system responsible for assembling the variable regions of immunoglobulin and T cell receptor genes from germline gene segments is the principal mechanism for generating a diverse set of antigen receptors. This V(D)J recombination reaction is essential for the proper development of the immune system. Impairment of recombination, as in mutant mice with severe combined immunodeficiency (scid) results in a virtual absence of mature B and T lymphocytes. In addition to its crucial role in lymphocyte development, V(D)J recombination is responsible for aberrant rearrangements that result in lymphoid neoplasms. Therefore, understanding the mechanism of V(D)J recombination is important for understanding both normal development of the immune system and for determining the molecular basis of oncogenic recombination events. Recent work has demonstrated that recombination involves double-strand DNA breaks at recombination signal sequences, resulting in flush breaks at signal ends and covalently sealed (hairpin) coding ends. These broken molecules are likely to be intermediates in recombination, and their identification supports a hairpin model for V(D)J recombination. This project will address several critical questions about the recombination mechanism using sensitive tools to characterize the structure of hairpin termini and products of hairpin opening generated in vivo in mouse thymocytes. Experiments will also test whether the hairpin opening reaction is qualitatively or quantitatively affected by the scid mutation, as previously proposed. Additional studies will probe early steps in the recombination mechanism, including the potential role of complex formation involving pairs of recombination signals, and will address the molecular basis of the requirement for a pair of signals of different spacer lengths (the """"""""12-23"""""""" rule). The functional organization of the recombination signals will also be examined. Proposed studies will illuminate basic features of the V(D)J recombination mechanism and its regulation, and will further define the molecular basis of the scid recombination defect. This knowledge will increase our understanding of how certain human DNA repair disorders deregulate V(D)J recombination, resulting in an increased frequency of oncogenic chromosome translocations, and will provide insight into the molecular basis of human immunodeficiencies.
Showing the most recent 10 out of 35 publications
