• ABNORMAL V(D)J RECOMBINATION IN ATAXIA TELANGIECTASIA MUTATED (ATM) DEFICIENT CD4/CD8 DOUBLE NEGATIVE THYMOCYTES

      Bowen, Steven Edward; Livak, Ferenc; Hodes, Richard J. (2012)
      The diversification of the lymphocyte antigen receptor occurs through the somatic recombination of discontinuous variable (V), diversity (D) and joining (J) gene segments at specific stages of lymphocyte development in a process known as V(D)J recombination. In T lymphocytes, four T cell receptor (TCR) loci rearrange at distinct developmental stages in the thymus. Recombination of the TCR β , TCR γ and TCR δ loci occurs during the CD4/CD8 double negative (DN) stage, and TCR α rearrangement occurs subsequently during the CD4/CD8 double positive (DP) stage. In DN thymocytes it has been unclear whether individual cells are capable of rearranging TCR β, TCR γ and TCR δ simultaneously or if there is a temporal separation of rearrangement at distinct loci. V(D)J recombination involves the transient induction and repair of DNA double strand breaks, which activate components of the cellular DNA damage response (DDR), mediated by the apical PI3-like kinase Ataxia Telangiectasia mutated (ATM). The DDR serves to physically stabilize broken DNA ends as well as transduce signals between the DSB and downstream effectors involved in cell cycle checkpoint activation, apoptosis, and DNA repair. Here we demonstrate that the absence of ATM impairs thymocyte development at the DN stage as a result of defective rearrangement at the TCR β locus. We also describe non-canonical TCR rearrangements that occur between TCR loci on different chromosomes (trans rearrangements) as a result of simultaneous recombination in DN thymocytes. We find that the frequency of trans rearrangement is dramatically increased in ATM-/- mice, and that cells carrying trans rearrangements can survive thymic selection and be found in the periphery. Furthermore we identify peripheral T cells in WT and ATM-/- mice that express a hybrid V γ -C β TCR chain on the cell surface encoded by a trans rearranged chromosome. Cells expressing the V γ -C β hybrid receptor are developmentally dependent on TCR α expression, CD4 or CD8 single positive, MHC restricted and capable of differentiating into memory T cells. This work provides insight into the molecular mechanism of TCR trans rearrangement, as well as the selection and function of rare but detectable TCR chains encoded by trans rearrangement in mouse T cells.