The interaction of hematopoietic growth factors with their cognate receptors results in the activation of one or more signal transduction pathways leading to changes in the expression of genes that regulate cell growth and differentiation. This proposal is directed toward the elucidation of those signaling elements triggered by the hormone erythropoietin and toward the characterization of the roles played by the early response proto-oncogenes c-myc and c-myb. The individual phosphoprotein involved in membrane and cytosolic signaling will be studied. Selective proteins will be cloned and sequenced and their intracellular concentrations will be modulated with antisense oligodeoxynucleotides and by transfection with constitutively expressed clones. Protein kinase C subclasses will be investigated. Erythroid cell subclasses will be quantified and their expression will be manipulated experimentally permitting the identification of subclass specific protein phosphorylations. The role of protein phosphatases 1 and 2A in erythropoietin signaling will be evaluated using the phosphatase inhibitor okadaic acid. A detailed study and comparison of erythropoietin and dimethyl sulfoxide signals to the proto-oncogene c-myc will be undertaken. Specifically, a comparison of the membrane and cytosolic phosphorylation events utilized by these two inducers of erythroleukemia cell differentiation will be made. The molecular mechanism of erythropoietin's signal to c-myb will be studied. The hypothesis that the signal to c-myb is mediated by a phosphatase dependent reaction operating on proteins of fos/jun (AP-1) complex will be evaluated. The results of these investigations will be directly relevant to our understanding of the control of red blood cell production and may provide insights into the development of new therapies for disorders of hematopoiesis.
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