The mpl ligand: molecular and cellular biology of the critical regulator of megakaryocyte development.

The development of blood cells requires the interplay of hematopoietic stem and progenitor cells, marrow stroma and polypeptide growth factors. Although many proteins are thought to support the expansion of megakaryocytic precursor cells (e.g., interleukin [IL]-3, c-kit ligand [KL]), identification of the late-acting, lineage-specific growth factor for platelet production, termed Thrombopoietin (Tpo), has remained elusive. Recently, characterization of the proto-oncogene c-mpl revealed structural homology with the hematopoietic cytokine receptor family. Based on the cell of origin of its cDNA, we hypothesized that the ligand for c-mpl might be identical with Tpo. Using BaF3 cells engineered to express c-mpl, we employed a functional expression strategy to clone its cDNA. At low concentrations, the recombinant protein supports the growth of megakaryocytic colonies, alone and together with either IL-3 or KL. For IL-3 this appears to be additive, for KL, true synergy was detected. At higher concentrations, the mpl ligand (ML) alone supported a near maximal number of very large megakaryocytic colonies. Using suspension cultures and human megakaryocytic cell lines, we have also shown that ML induces the terminal differentiation of megakaryocytes by enhancing polyploidization and surface membrane expression of GPIb and IIb/IIIa. Moreover, the development of megakaryocytes in vitro appears to be absolutely dependent on the presence of ML. Following receptor engagement, ML induces tyrosine phosphorylation of a number of membrane associated kinases and adaptor molecules, including SHC, JAK2, PLC-gamma and the mpl receptor itself.(ABSTRACT TRUNCATED AT 250 WORDS)