Functional Interaction of c-Kit Receptor and Interleukin-3 Receptor.

Hematopoiesis is regulated by a number of growth factors and cytokines, among which stem cell factor (SCF) plays a critical role for normal hematopoiesis. An important feature of SCF is its strong capacity to synergize with the cytokines including interleukin-3(IL-3), granulocyte-marcrophage colony-stimulation factor (GM-CSF) and erythropoietin to regulate proliferation and differentiations of hematopoietic progenitor cells. The mouse hematopoietic progenitor cell line EML contains two populations of lineage negative cells, distinguished by the expression or absence of CD34. The two sub-populations express similar levels of the SCF receptor c-kit, but only the CD34+ population replicates in the presence of SCF alone, while the CD34 negative population replicates in the presence of IL3 but this replication is synergistically stimulated by the addition of SCF. Our previous studies indicated that synergistic proliferation of the mouse hematopoietic progenitor cell line EML induced by SCF in combination with IL-3 was probably mediated by the synergistic increases in tyrosine phosphorylation of c-Kit and β-chain of IL-3 receptor, which is shared with GM-CSF receptor and interleukin-5 receptor. Conversely, the decreased response of c-kit to SCF alone in the CD34 negative population may be a result of the increased expression of IL3 receptor in these cells. Trans-phosphorylation between c-Kit and the β-chain of IL-3 receptor was also observed in EML cells. In studies of the molecular mechanism behind the functional interactions between SCF and IL-3 we found that c-Kit and IL-3 receptor β-chain form a complex in EML cells. Antibody Ab-1 specific against the Ig-like domain 4 in the extracellular region of c-Kit blocked not only cell proliferation induced by SCF but also synergistic proliferation induced by SCF plus IL-3. Consistent with this finding, Ab-1 inhibited phosphorylation of c-Kit induced by SCF or IL-3, and synergistic phosphorylation of c-kit induced by SCF plus IL-3 respectively. SCF and IL-3 in combination synergistically activated the protein kinases Akt and Erks, the downstream mediators of c-Kit and IL-3R. Phosphorylation of Erks is independent on the phosphatidylinositol 3-kinase (PI3-kinase) pathway in EML cells. These data suggested the possibility that the synergistic growth of EML cells induced by SCF in combination with IL-3 was mediated by multiple mechanisms that could include trans- and synergistic phosphorylation of c-Kit and the β-chain of IL-3 receptor, PI3-kinase dependent synergistic phosphorylation of Akt and PI3-kinase independent synergistic phosphorylation of Erks. The trans- and synergistic phosphorylation of c-Kit and β-chain of IL-3 receptor was mediated by physical interaction of two receptors.