T cell receptor (TCR) locus enhancer identity and position are critical for the assembly of TCR and variable region genes

T cell receptor (TCR) and variable region genes are assembled from germ-line gene segments located in a single chromosomal locus in which TCR segments are situated between TCR segments. The TCR enhancer (E ) located at the 3 end of the TCR locus functions over a long chromosomal distance to promote TCR rearrangement and maximal TCR expression; whereas the TCR enhancer (E ) is located among the TCR segments and functions with additional element(s) to mediate TCR rearrangement. We used gene-targeted mutation to evaluate whether the identity of E and the position of E are critical for the developmental stage-specific assembly of TCR and variable region genes. Specific replacement of E with E , the core E element (E C), or the Ig heavy chain intronic enhancer (iE ), all of which promote accessibility in the context of transgenic V(D)J recombination substrates, did not promote a significant level of TCR rearrangement beyond that observed in the absence of E . Therefore, the identity and full complement of E -binding sites are critical for promoting accessibility within the TCR locus. In the absence of the endogenous E element, specific replacement of E with E also did not promote TCR rearrangement. However, deletion of intervening TCR locus sequences to restore the inserted E to its normal chromosomal position relative to 5 sequences rescued TCR rearrangement. Therefore, unlike E , E lacks ability to function over the large intervening TCR locus and or E function requires proximity to additional upstream element(s) to promote TCR accessibility.