Heterogeneity of N insertion capacity in fetal hematopoietic stem cells

TCR gene rearrangement is strictly regulated during mouse ontogeny. The V-(D)-J junctions of alphabeta and gammadelta TCR transcripts expressed in the adult thymus are more highly diverse than those in the fetal thymus. We previously showed that adult hematopoietic stem cells (HSC) have a higher capacity to insert N nucleotides into Vgamma4 TCR transcripts than fetal HSC and that the level of N nucleotide insertion is determined, at least in part, at the level of HSC. To analyze this developmental change of HSC at the single cell level, we investigated N nucleotide insertions in three TCR transcripts (Vgamma4, Vgamma2 and Vbeta8) 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 Vgamma transcripts (0-78%). On the other hand, the level of N insertions was similarly regulated in Vgamma4, Vgamma2, and Vbeta8 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. After 3 weeks culture with a stromal cell line, fetal HSC showed higher levels of N insertion capacity than before culture. This result and the presence of HSC with intermediate N insertion capacity support the hypothesis that the developmental potential of individual HSC gradually changes from fetal to adult type in one stem cell lineage.