This research project concerns the relevance to hematopoietic differentiation and myeloproliferative disease of a 'new' Mr 66 000 protein that we have discovered in erythrocyte membranes from all tested patients with polycythemia vera, PV, and a patient with chronic myelogenous leukemia. We will evaluate the significance of this by measuring its incidence in other conditions suspected of being disorders in stem cell proliferation, myelofibrosis, thrombocythemia, paroxysmal nocturnal hemoglobinuria and chronic granulocytic leukemia, and analysis of the synthesis of 'PV protein' stem cells from patients with PV, other myeloproliferative disorders and normal individuals in vitro. For the latter we will employ labeling with 35S-methionine and 14C-oligosaccharide precursors and measure possible changes in the production of 'PV protein' by normal progenitors in response to erythropoietin. As controls we will also evaluate secondary erythremia and iron deficiency states. To determine the cellular distribution of 'PV protein', we will assay membranes from PV and control neutrophils, platelets as well as from PV reticulocytes and mature erythrocytes of varying ages. Assay of the protein will be by bidimensional isoelectric focusing/SDS PAGE and by immunochemical methods. We will also sharpen estimates of Mr, and pI, and characterize the immunoprecipitated, SDS PAGE-separated protein by tryptic peptide mapping.
