HETEROGENEITY OF N-INSERTION CAPACITY IN FETAL HEMATOPOIETIC STEM-CELLS

TCR gene rearrangement is strictly regulated during mouse ontogeny. Th e V-(D)-J junctions of alpha beta and gamma delta TCR transcripts expr essed in the adult thymus are more highly diverse than those in the fe tal thymus, We previously showed that adult hematopoietic stem cells ( HSC) have a higher capacity to insert N nucleotides into V(gamma)4 TCR transcripts than fetal HSC and that the level of N nucleotide inserti on is determined, at least in part, at the level of HSC. To analyze th is developmental change of HSC at the single cell level, we investigat ed N nucleotide insertions in three TCR transcripts (V(gamma)4, V(gamm a)2 and V(beta)8) derived from limiting numbers of fetal liver HSC by fetal thymic organ culture, Eight day-14 fetal liver HSC clones showed various levels of N nucleotide insertions in V(gamma)4 transcripts (0 -78%). On the other hand, the level of N insertions was similarly regu lated in V(gamma)4, V(gamma)2 and V(beta)8 TCR transcripts in a clone- specific way, These results suggested that the level of N insertion is programmed at the level of single HSC and that fetal liver contains a heterogeneous population of HSC in terms of N insertion capacity. Aft er 3 weeks culture with a stromal cell line, fetal HSC showed higher l evels of N insertion capacity than before culture. This result and the presence of HSC with intermediate N insertion capacity support the hy pothesis that the developmental potential of individual HSC gradually changes from fetal to adult type in one stem cell lineage.