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Induction of the Receptor for Erythropoietin in Murine Erythroleukemia Cells after Dimethyl Sulfoxide Treatment¹

The Third Department of Internal Medicine, Faculty of Medicine, University of Tokyo, 7-3-1 Hongo, Tokyo, Japan 113

Biologically active ¹²⁵I-labeled human recombinant erythropoictin (EPO) was used to demonstrate specific receptors for this erythroid-specific hemopoietic growth factor on the cell surface of murine erythroleukemia cell clone B8. The binding of radioiodinated EPO to these cells was time and temperature dependent, specific, saturable, and reversible. During erythroid differentiation by dimethyl sulfoxide, B8 cells displayed a rapid and marked increase in the amount of specific ¹²⁵I-EPO binding before the appearance of hemoglobin-containing cells. Scatchard analysis of the saturation binding data revealed that B8 cells had a single class and low number (350 to 650) of EPO receptors per cell with an apparent K₄ of 1.2 to 1.4 nM. In addition, the number of EPO receptors on B8 cells was increased twice by induction with DMSO for 1 day, but the binding affinity of EPO toward its receptors did not change significantly. Affinity cross-linking experiments with disuccinimidyl suberate demonstrated two radiolabeled components with apparent molecular weights of 145,000 and 130,000 under both reducing and nonreducing conditions. Labeling of the two components was inhibited by incubation of cells with unlabeled EPO. These results suggest that some murine erythroleukemia cells potentially express EPO receptors as a differentiation marker of erythroid lineage, which contain two polypeptides with molecular weights of 109,000 and 94,000.

¹ This work was supported by a research grant for intractable diseases from the Ministry of Health and Welfare of Japan, and grants-in-aid from the Ministry of Education, Science, and Culture of Japan.

² To whom requests for reprints should be addressed.

Received 7/27/87. Revised 11/11/87. Accepted 12/23/87.