Elucidating the role of cis-regulatory elements in mediating V(D)J recombination

Harvard Medical School, Boston MA

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Abstract

? DESCRIPTION (provided by applicant): Adaptive immunity is dependent on the ability of B and T cell receptors (BCR; TCR) to recognize and bind foreign substances within the body. Primary diversity in these BCR and TCR repertoires is dependent on the gene rearrangement process termed V(D)J recombination, in which controlled rearrangements of V, D, and J segments occur across large genomic distances in developing lymphocytes to produce genetically unique BCRs or TCRs. Importantly, deregulation of V(D)J recombination can underlie immunodeficiency and autoimmunity, and also lead to oncogenic translocations. At the Immunoglobulin heavy chain (IgH) locus, V(D)J recombination is tightly regulated. This regulation can be summarized into the following four properties: 1. Lineage specificity - Complete VHDJH joins from VH to DJH rearrangements are only observed in B cells and not T cells 2. Stage specificity - IgH rearrangements in progenitor B cells prior to light chain rearrangements, which occur in pre-B cells. 3. Order - D to JH rearrangements occur on both IgH alleles before joining of a VH to DJH rearrangement. VHs do not join to unrearranged Ds. 4. Feedback regulation - A productive VHDJH rearrangement on one allele will produce a heavy chain that feeds back to prevent further VH to DJH rearrangements on the second allele, if it is still in DJH configuration. The Alt lab has demonstrated that a 4kb region within the 100b IgH VH-D intergenic region, termed Intergenic Control Region 1 (IGCR1), contains two CTCF-binding elements (CBEs) that are essential in cis for ordered, lineage-specific, and feedback regulated assembly of VH, D, and JH segments. These functions of IGCR1 diversify primary antibody repertoires by preventing premature rearrangements of VH81X. IGCR1 also forms chromatin loops with other enhancer and CBE sites within the locus, but a specific mechanism for how these loops or other elements may participate in IGCR1 functions is not known. Thus, my proposal aims to (1) elucidate individual roles of the CBEs within IGCR1 and a set of ten CBEs at the 3' end of the IgH locus in V(D)J recombination control, and (2) test for transcription- and position-dependent aspects of IGCR1 functions. My proposed project should provide new insights into the interplay of genetic and epigenetic factors that that govern V(D)J recombination, which also will translate into a broader understanding of gene regulation.