c-kit ligand stimulates tyrosine phosphorylation of a similar pattern of phosphotyrosyl proteins in primary primitive normal hematopoietic progenitors that are constitutively phosphorylated in comparable primitive progenitors in chronic phase chronic myelogenous leukemia

Characteristic of Philadelphia (Ph)⁺ chronic myelogenous leukemia (CML) is the presence of the chimeric BCR/ABL (p210) protein possessing elevated protein tyrosine kinase activity relative to the normal c-abl tyrosine kinase. Our previous studies demonstrated subtle differences in the growth, phenotypic and morphologic characteristics of the most primitive subpopulations of primary lin^- Ph⁺ chronic phase CML blasts and comparable primary lin normal blasts. Recently, in comparing proteins phosphorylated on tyrosine in these cell populations, we reported a prominent 62 kDa phosphotyrosyl (P-tyr) protein constitutively present in primary primitive lin^- CML chronic phase blasts which was virtually undetectable in primary primitive lin^- normal blasts. In the present studies, we demonstrate that this P-tyr p62 from primary primitive lin^- chronic phase CML blasts co-immunoprecipitates with ras-GAP. Furthermore, in addition to the p210 protein, we show in whole cell lysates the presence of other clearly consistent but less prominent P-tyr proteins with molecular weights of ∼155, 140, 110, 55 and 45 kDa as well as more minor P-tyr proteins of ∼190, 85, 52, 42 and 39 kDa constitutively present in primary primitive lin^- chronic phase CML blasts. In analyzing proteins tyrosine phosphorylated in primary primitive lin^- normal blasts in response to various hematopoietic growth factors, we found a striking similarity in the phosphorylation of four major (∼140, 110, 62 and 56 kDa) and three minor (∼51, 45 and 42 kDa) P-tyr proteins after stimulation with c-kit ligand and the P-tyr proteins constitutively phosphorylated in primary primitive lin chronic phase CML blasts. Other growth factors tested (ie GM-CSF, G-CSF, IL-3, FLT3 ligand and EPO) were much less active or stimulated phosphorylation of other proteins. It is provocative that at least seven proteins rapidly and transiently phosphorylated on tyrosine in the c-kit ligand signal transduction pathway in lin^- normal blasts may be constitutive substrates for the p210 activated tyrosine kinase in comparable lin^- chronic phase CML blasts. In addition, it is intriguing that some of the biological effects on hematopoietic progenitors attributed to the c-kit ligand may be similar to some of the observed biological consequences of the p210 protein, including survival and expansion of a more mature stem cell population, probably at the time of lineage commitment rather than at the level of the earliest self-renewing stem cell.