scTCR-seq and HTS reveal a special novel TRBD2-TRBJ1 rearrangement in mammalian TRB CDR3 repertoire.

Mammalian T cell receptor (TCR) beta-chain (TRB) V-D-J rearrangement mainly follows the "12/23 rule", and the "D-J rearrangement preceding the V-(D-J) rearrangement". Owing to the physical position of the D-J-C cluster in the TRB locus, the TRBD2 (D2) gene cannot directly perform inversional rearrangement or deletional/loop-out rearrangement with the TRBJ1 (J1) gene. Our previous studies revealed a single reverse TRBV30 (TRBV31 in mice) gene in the mammalian TRB locus, which can cause indirect rearrangement of the D2 gene and J1 gene; however, the mechanism and proportion involved in germline gene rearrangement are unknown. We obtained TRB CDR3 repertoires of thymus and peripheral tissues from humans and mice by HTS and scTCR-seq and found that 14% of the rearrangements in which the D2 gene is involved are D2-J1 rearrangements (D2-J2 rearrangements account for approximately 86%). The mechanism is that the reverse V30 gene preferentially performs inversional rearrangement with the D2 gene (V30-D2), leading to V30-D2-J1 rearrangement in humans, or the reverse V30 gene preferentially performs inversional rearrangement with the D1 gene (V30-D1), allowing the forward V genes (Vx) to perform Vx-D2-J1 rearrangement. We further found that D2-J1 rearrangements were present in more than 24% and more than 15% of the D2 gene rearrangements in rhesus monkeys and bats, respectively. Moreover, in bovine containing D1J1C1, D3J3C3, and D2J2C2 clusters, more than 11% D3-J1 and D2-J1 rearrangements and more than 22% D2-J3 rearrangements were found. This study provides a new perspective and feasible solution for further research on the significance of the special V-D-J recombination pattern in the mammalian TRB locus and the CDR3 repertoire formed by D2-J1 rearrangement.