Orchestrating T-cell receptor α gene assembly through changes in chromatin structure and organization

Abstract

V(D)J recombination is regulated through changes in chromatin structure that allow recombinase proteins access to recombination signal sequences and through changes in three-dimensional chromatin organization that bring pairs of distant recombination signal sequences into proximity. The Tcra/Tcrd locus is complex and undergoes distinct recombination programs in double negative and double positive thymocytes that lead to the assembly of Tcrd and Tcra genes, respectively. Our studies provide insights into how locus chromatin structure is regulated and how changes in locus chromatin structure can target and then retarget the recombinase to create developmental progressions of recombination events. Our studies also reveal distinct locus conformations in double negative and double positive thymocytes and suggest how these conformations may support the distinct recombination programs in the two compartments.

Figure 1. Organization of the Tcra/Tcrd locus

Gene segments are depicted by filled rectangles, enhancers are depicted by filled ovals, and prominent promoters in the Jα array are depicted by bent arrows. The commonly used Vδ gene segments (TRAV15/DV6 family, TRDV2-2, TRDV4, TRDV5) are identified. Note that TRDV4 rearranges preferentially in fetal thymocytes.

Figure 2. Regulation of Jα chromatin accessibility

Wild-type and genetically manipulated versions of the Jα array are depicted. ΔTEA and ΔTEAΔJ49 carry deletions of the TEA and J49 promoters. TEA-T and 56R carry insertions of a transcription terminator downstream of TEA or Jα56, respectively. HYα carries an insertion of a TRAV17-Jα57 rearrangement. Large arrows identify major promoters (TEA, J49). Small arrows identify minor or cryptic promoter activity. Shaded portions of the ΔTEA,ΔTEAΔJ49, TEA-T and 56R alleles identify areas of diminished chromatin modifications and disrupted Vα-to-Jα recombination.

Figure 3. Tcra/Tcrd locus conformational states

The diagram depicts the relatively extended configuration of the locus in B cells, the fully contracted configuration in DN thymocytes, and the unique 3’ contracted and 5’ decontracted configuration in DP thymocytes. The V array is segregated into distal (5’), central, and proximal (3’) regions.