DIFFERENT DEVELOPMENTAL ARREST POINTS IN RAG-2- - AND SCID THYMOCYTESON 2 GENETIC BACKGROUNDS - DEVELOPMENTAL CHOICES AND CELL-DEATH MECHANISMS BEFORE TCR GENE REARRANGEMENT/

To analyze the early development of T cell precursors in the absence o f TCR gene rearrangement, recombinase-activating gene-deficient (RAG-2 -/-) thymocytes were compared with thymocytes from SCID mice on the C .B-17 (BALB) and B6 genetic backgrounds. RAG-2 -/- thymocytes accumula te as quiescent cells with a heat-stable Ag (HSA)-positive CD25(+)CD44 (-)c-kit(low) phenotype, resembling normal cells just before selection for functional TCR beta-chain expression. CD44 and c-kit progressivel y down-regulate in the HSA(+) subset, providing a background-independe nt and TCR-independent developmental clock. On this basis, compared wi th RAG-2 -/- thymocytes, SCID thymocytes 1) arrest at more heterogeneo us, and generally earlier, stages; 2) accumulate to lower overall cell numbers; and 3) maintain higher populations of cycling and activated G(1) cells, showing both increased responsiveness and increased cell d eath. B6-SCID thymocytes appear to die particularly early, Low levels of Fas were observed on ''advanced'' HSA(+) SCID thymocytes but not on any RAG-2 -/- thymocytes, suggesting a potential difference in activa tion state or mechanism of death. In both RAG-2 -/- and SCID thymocyte s, there are also two discrete subsets of HSA(low)CD25(-)CD44(+)c-kit( +) cells: a Sca-1(+)CD44(++)CCD122-NK1.1(-) putative progenitor subset and an NK-like Sca-1(-)CD44(+(+))CD122(+)NK1.1(+) subset. The absolut e cell numbers in these HSA(low) subsets and the extent of NK cell dif ferentiation, measured by perforin expression, are nearly constant in all the mutant strains analyzed, in contrast to the HSA(+)CD25(+) popu lation, which was expanded in the RAG-2 -/-. Thus, the SCID thymocytes appear to undergo a normal generation but a premature death as compar ed with the RAG-2 -/- thymocytes.