Alterations of the phosphoinositide 3-kinase and mitogen-activated proteinkinase signaling pathways in the erythropoietin-independent Spi-1/PU.1 transgenic proerythroblasts

During the cell transformation processes leading to erythroleukemia, erythr oid progenitors often become erythropoietin (Epo)-independent for their pro liferation. The biochemical events that could lead an erythroleukemic cell to growth factor-independence were investigated using spi-1 transgenic poer ythroblasts. Spi-1/ PU.1 is a myeloid and B-cell transcription factor of th e ETS family and is activated by insertional mutagenesis during Friend eryt hroleukemia. Its overexpression in proerythroblasts induces their different iation arrest without altering their erythropoietin requirement for prolife ration (HS1 cells). At a later step, genetic alterations most probably occu r allowing spi-1 transgenic poerythroblasts to proliferate in the absence o f erythropoietin (HS2 cells). The signaling transduction pathways in HS1 an d HS2 proerythroblasts were analyzed. The authors have previously shown tha t the Jak/STAT pathway was not activated in Epo-independent cells, but rema ined sensitive to Epo stimulation. In the present study, it is shown that t he Epo-independent proliferation of HS2 cells requires active phosphoinosit ide 3-kinase (PI3K) and mitogen-activated protein kinase (MAPK) pathways. I n these cells, PI3K was constitutively associated with the molecular adapte rs Grb2 and Gab1, and with the phosphatases SHP-2 and SHIP. Moreover, PI3K activity was correlated with the constitutive phosphorylation of serine-thr eonine protein kinase (AKT) in HS2 cells. Lastly, a constitutive activation of the MAPKs extracellular signal-regulated kinases (ERK1/2) in HS2 cells was observed that occurs in a PI3K-independent manner, but depends strictly on the activity of the protein kinase C (PKC). These results suggest that constitutive activations of PI3K/AKT and PKC/MAPK pathways can act in syner gy to lead a proerythroblast to proliferate without Epo. (C) 2001 by The Am erican Society of Hematology.