CELL-CYCLE CONTROL OF C-KIT(-7R(+) B-PRECURSOR CELLS BY 2 DISTINCT SIGNALS DERIVED FROM IL-7 RECEPTOR AND C-KIT IN A FULLY DEFINED MEDIUM()IL)

An important goal for the investigation of the proliferation of mammal ian cells is to establish a fully defined condition for culturing them in vitro. Here, we report establishment of a fully defined culture co ndition that supports the primary culture of normal c-kit(+)IL-7 recep tor (IL-7R)(+) B precursor cells without the aid of stromal cell lines . This defined culture condition contains IL-7, the ligand for c-kit, transferrin, insulin, and bovine serum albumin as protein components. By using the cell lines derived from RAG2(-/-) mice, which do not diff erentiate into c-kit(-) stage, we have evaluated the role of each prot ein in the cell cycle progression of c-kit(+)IL-7R(+) B precursor cell s. Since B precursor cells can grow without insulin, c-kit remains a s ole functional receptor tyrosine kinase for their growth. While both c -kit ligand (KL) and IL-7 are the requisite molecules for sustained pr oliferation of B precursor cells, each molecule plays distinct roles. IL-7 starvation results in prompt arrest of the cells at G1. An accumu lation of the cells in the mitotic phase was also detected. Thus, the major role of IL-7 is to regulate the G1/S transition and the process of cytokinesis of B precursor cells. Although prolonged KL starvation over 48 h resulted in accumulation of G1 cells, its effect could not b e detected within 24 h, which is long enough for all the cells to comp lete one cell cycle. This suggests that KL might be involved in the ce ll cycle progression of B precursor cells in a manner that its signal could still be effective in the one or two cell cycles that follow. Al though molecular nature of the signals underlying the present observat ion awaits future investigation, the method described in this report w ould provide a useful model system for investigating the signaling pat hways that are involved in the cell cycle progression of B precursor c ells.