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The TCR Enhancer Is Dispensable for the Expression of Rearranged TCR Genes in Thymic DN2/DN3 Populations but Not at Later Stages

Department of Cellular Immunology, Max-Planck-Institute of Immunobiology, Freiburg, Germany

The E enhancer has been shown to be dispensable for germline transcription of nonrearranged TCR segments but appears to be required for TCR V to DJ rearrangement. E dependency of the subsequent expression of VDJ-rearranged TCR genes in thymic subpopulations has so far not been analyzed. We generated transgenic mice, using a V8.2D1J1.3-rearranged TCR bacterial artificial chromosome, which lacked E, and monitored transgene expression by flow cytometry using V-specific mAbs and an IRES-eGFP reporter. Transgene expression was found in double negative (DN)2 and DN3 but not at later stages of thymopoesis. There was no toxicity associated with the transgene given that apoptosis in DN3, DN4 was not increased, and the number of DN4 cells generated from DN3 cells in reaggregate thymic organ cultures was not diminished. The transgenic TCR gave rise to a pre-TCR, as suggested by its ability to suppress endogenous TCR rearrangement, to facilitate -selection on a TCR-deficient background and to inhibit T cell lineage development. The results suggest that the V8.2 promoter is sufficient to drive expression of rearranged TCR VDJ genes E independently in DN2/DN3 but not at later stages.

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