DNA recombination and somatic hypermutation are hallmarks of the immunoglobulin (Ig) loci in developing and antigen-reactive B lymphocytes. B lymphocytes must assemble the genes encoding Ig chains, and this process begins with VH gene assembly. Somatic hypermutation (SHM), together with clonal selection, insures the development of high affinity antibodies to invading pathogens. Class-switch recombination (CSR) makes possible the development of antigen-reactive Ig's of multiple isotypes so that these Ig's can gain access to antigen wherever it resides and eliminate it through any of a number of mechanisms. ? ? These genome-damaging processes must be strictly controlled so that they act only on the Ig loci, where they are needed for normal locus function. This regulatory responsibility appears to fall on a series of cis- acting elements found within and around the Ig heavy (IgH) and light chain (IgL) loci. It has become increasingly clear that improper targeting of somatic hypermutation and class-switch recombination is a contributing factor, if not the initiating factor, in numerous neoplasms arising within the B lymphoid compartment. Not only does mis-targeting of the DNA lesions lead to undesirable chromosome translocations, but also, transposition of the Ig regulatory sequences to sites adjacent to proto-oncogenes leads to unwanted gene activity (reviewed in 36). Further study of the regulatory elements in the IgH locus, therefore, must be an essential component of any effort aimed at treating and/or preventing B-lineage cancers, as well as of efforts to harness the power of this system to more effectively combat infection. ? ? In this proposal, we seek to more precisely delineate the regulatory sequences within the IgH locus that regulate VH gene assembly (in particular, D-J joining), CSR, and SHM. We have developed bacterial artificial chromosomes (BACs) that carry an assembled VH gene and the whole of the IgH locus downstream. These BACs have been designed so that after they are integrated into the genome of transgenic animals, individual (and pairs of) elements within the 3'lgH regulatory region can be deleted at will. This experimental system will be used to further study a system of control elements that we, and others, have shown to have substantial functional redundancy. In addition, we have recently developed a mouse that lacks E mu (intronic enhancer) but carries a fully-assembled VH gene. This experimental system allows us not only to assess E mu's essential functions post-V(D)J assembly, but also its role in allelic exclusion. An understanding of the latter will contribute to models relating allelic exclusion to protection from autoimmunity. ? ? ?
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