The significance of a population in mouse bone marrow of lineage-negative (Lin-), Sca1-positive, c-kit-negative (LSK-) cells, which is reported to be devoid of long-term repopulation capacity or myeloid potential, is unknown. In this study, we show that the LSK- population is composed of several subsets defined by the expression of flt3, CD25, and IL-7Rα. The first subset was CD25- and more than 90% expressed either flt3, IL-7Rα, or both. The CD25-LSK - population had T cell, B cell, and NK cell potential in vivo, and most of this activity was localized in the flt3+ subset, irrespective of the expression of IL-7Rα. Although lymphoid potential of flt3 +LSK- cells in vivo was 3-fold lower than that of lin -Sca1lowkitlowIL7Rα+ common lymphoid progenitors (CLPs), their cloning efficiency in vitro was 10-fold lower than that of CLPs. Furthermore, although the myeloid potential of flt3 +LSK- cells was 10-fold lower than that of CLPs in the absence of M-CSF, the relative myeloid potential of both populations was similar in its presence. These observations suggest differential growth factor requirements of both populations. The second subset of LSK- cells was homogeneously CD25+flt3-IL7Rα+ and could be generated from both CD25-LSK- cells and from CLPs, but did not engraft in immunodeficient Rag1-/- or Rag1 -/-γc-/- hosts. This population, of which the significance is unclear, was increased in Rag1-/- mice and in old mice. Thus, the LSK- population is phenotypically and functionally heterogeneous and contains early lymphoid-committed precursors. Our findings imply that the early stages of lymphoid commitment are more complex than was thus far assumed.