Dysregulated expression of pre-Talpha reveals the opposite effects of pre-TCR at successive stages of T cell development.

The pre-TCR complex (TCRbeta-pre-TCRalpha chain (pTalpha)), first expressed in a fraction of CD8(-)4(-)CD44(-)25(+) (DN3) cells, is believed to facilitate or enable an efficient transition from the CD8(-)4(-) double-negative (DN) to the CD8(+)4(+) double-positive (DP) developmental stage. Subsequent to pre-TCR expression, DN3 thymocytes receive survival, proliferation, and differentiation signals, although it is still unclear which of these outcomes are directly induced by the pre-TCR. To address this issue, we generated mice bearing a range of pTalpha transgene copy number under the transcriptional control of the p56(lck) proximal promoter. All lines exhibited increased DN3 cycling, accelerated DN3/4 transition, and improved DN4 survival. However, the high copy number lines also showed a selective reduction in thymic cellularity due to increased apoptosis of DP thymocytes, which could be reversed by the ectopic expression of Bcl-2. Our results suggest that transgenic pTalpha likely caused apoptosis of DP thymocytes due to competitive decrease in surface TCRalphabeta formation. These results highlight the critical importance of precise temporal and stoichiometric regulation of pre-TCR and TCR component expression.

Mapping precursor movement through the postnatal thymus reveals specific microenvironments supporting defined stages of early lymphoid development.

Cellular differentiation is a complex process involving integrated signals for lineage specification, proliferation, endowment of functional capacity, and survival or cell death. During embryogenesis, spatially discrete environments regulating these processes are established during the growth of tissue mass, a process that also results in temporal separation of developmental events. In tissues that undergo steady-state postnatal differentiation, another means for inducing spatial and temporal separation of developmental cues must be established. Here we show that in the postnatal thymus, this is achieved by inducing blood-borne precursors to enter the organ in a narrow region of the perimedullary cortex, followed by outward migration across the cortex before accumulation in the subcapsular zone. Notably, blood precursors do not transmigrate the cortex in an undifferentiated state, but rather undergo progressive developmental changes during this process, such that defined precursor stages appear in distinct cortical regions. Identification of these cortical regions, together with existing knowledge regarding the genetic potential of the corresponding lymphoid precursors, sets operational boundaries for stromal environments that are likely to induce these differentiative events. We conclude that active cell migration between morphologically similar but functionally distinct stromal regions is an integral component regulating differentiation and homeostasis in the steady-state thymus.

Cellular interactions in the thymus regulate the protein kinase C signaling pathway.

We provide evidence that thymocytes receive signals from the thymic microenvironment which regulate the protein kinase C (PKC) signaling pathway. Thus, phorbol 12-myristate 13-acetate (PMA) causes a PKC-dependent down-regulation of CD4 expression and induces apoptosis in isolated thymocytes but has little effect on thymocytes maintained within intact thymic lobes or in reaggregate lobes containing purified thymocytes with either thymic or non-thymic stromal cells. Moreover, compact pellets of thymocytes alone are protected from the effects of PMA. This protection is maintained when the compacted thymocytes are rigorously depleted of MHC class II-expressing cells. We conclude that signals arising from thymocyte-thymocyte contact control the utilization of the PKC cascade. These observations have implications for thymocyte signaling in general as well as for the interpretation of studies carried out on thymocyte suspensions.