Promoter-dependent Epigenetic Regulation of V(D)J Recombination

Human development requires the stable and heritable activation of discrete genome regions in developing tissue without altering DMA sequence integrity. The developmental mechanisms by which this epigenetic regulation is imposed are only just coming to light. One startling example of epigenetic regulation occurs in developing lymphocytes which assemble antigen receptor genes from pools of related gene segments via a series of somatic rearrangements termed V(D)J recombination. Rearrangement of each antigen receptor gene is restricted to a discrete lineage and stage of lymphopoiesis. For example, developing T cells assemble genes for the T cell receptor (TCR) by first rearranging the beta chain gene in a two-step process involving D-to-J, then V-to-DJ recombination. TCRbeta recombination occurs early in T cell development, and is necessary for developmental progression and initiation of TCRalpha assembly. Such tight control is central to production of mature lymphocytes and safeguards the integrity of the genome. Indeed, mistargeting of recombination has been linked to a variety of leukemias. Gene segment targeting requires modulations in their chromatin accessibility by associated transcriptional enhancers and promoters. However, it remains unclear how these elements coordinate differential accessibility of gene segments within a single locus. This proposal tests the hypothesis that developmental variations in the activation and strength of individual promoters direct differential accessibility of associated gene segments. Using TCRbeta as a model, we propose to examine the roles of promoters in epigenetic regulation of recombinational accessibility by (1) dissecting promoter contributions to differential recombination of Dbetal and Dbeta2; (2) defining promoter contributions to activating V recombination; and (3) defining the roles of V and D RS/promoter combinations within the locus in limiting inappropriate D rearrangements. Results promise insight into the general mechanisms by which cis-acting elements direct epigenetic control of gene activation. V(D)J recombination allows us to generate a robust immune system. But altering DMA can be dangerous, leading to cancer formation. By understanding the mechanisms that target recombination at specific DMA sites, we will gain insight into to pathways that lead to inappropriate, cancer-causing rearrangements. Such insights will be important for identifying potential cancers before they develop.