Long-Distance Regulation of Fetal V

Murine Tcra and Tcrd gene segments are organized into a single genetic locus (Tcra/Tcrd locus) that undergoes V(D)J recombi-nation in CD4 2 CD8 2 double-negative (DN) thymocytes to assemble Tcrd genes and in CD4 + CD8 + double-positive thymocytes to assemble Tcra genes. Recombination events are regulated by two developmental stage-specific enhancers, E d and E a. Effects of E a on Trca/Tcrd locus chromatin have been well documented, but effects of E d have not. In this regard, E a acts over long distances to activate many V a and J a segments for recombination in double-positive thymocytes. However, in DN thymocytes, it is unclear whether E d functions over long distances to regulate V d gene segments or functions only locally to regulate D d and J d gene segments. In this study, we analyzed germline transcription, histone modifications, and recombination on wild-type and E d-deficient alleles in adult and fetal thymocytes. We found that E d functions as a local enhancer whose influence is limited to no more than ∼10 kb in either direction (including D d , J d , and TRDV5 gene segments) in adult DN thymocytes. However, we identified a unique long-distance role for E d promoting accessibility and recombination of fetal V d gene segment TRDV4, over a distance of 55 kb, in fetal thymocytes. TRDV4 recombination is specifically repressed in adult thymocytes. We found that this repression is enforced by a developmentally regulated loss of histone acetylation. Constitutively high levels of a suppressive modification, histone H3 lysine 9 dimethylation, may contribute to repression as well. T he development of ab and gd T lymphocytes depends on the somatic assembly of TCR genes by V(D)J re-combination (1). Among the four TCR genes, Tcrd and Tcra are uniquely organized into a single, complex genetic locus (the Tcra/Tcrd locus) that spans 1.6 megabases of murine chromosome 14 (2). Tcra/Tcrd locus recombination events occur according to a strict developmental program during thymocyte maturation, with Tcrd genes assembled in CD4 2 CD8 2 double-negative (DN) thymocytes and Tcra genes assembled in those thymocytes that progress to the CD4 + CD8 + double-positive (DP) stage. Developmental programming is thought to be mediated by changes in chromatin structure that make certain recombination signal sequences (RSSs) accessible to the recombination activating gene complex and by changes in locus organization that allow specific pairs of RSSs to undergo synapsis for recombination …