FUNCTIONAL ERYTHROPOIETIN RECEPTOR OF THE CELLS WITH NEURAL CHARACTERISTICS - COMPARISON WITH RECEPTOR PROPERTIES OF ERYTHROID-CELLS

Radioiodinated erythropoietin (Epo) was bound specifically to the cell s of two non-erythroid clonal lines, PC12 and SN6, which expressed neu ronal characteristics. The binding was time-, cell number-, and dose-d ependent and was reversible. Although the cloned Epo receptor from PC1 2 cells (derived from rat adrenal medulla) was identical to that from rat erythroid cells, significant differences in the ligand binding pro perties between two cell lineages were found; 1) PC12 cells had a sing le class of binding sites with very low affinity (K(d) = 16 nM), where as erythroid cells had two classes of binding sites with different aff inities (K(d) = 95 pM for high affinity sites and 1.9 nM for low affin ity sites), and 2) cross-linking experiments revealed one cross-linked product of 105 kDa for PC12 cells and two products of 140 and 120 kDa for erythroid cells. Taken together with additional results, the pres ence of a putative accessory protein(s) that may alter the ligand bind ing affinity through interaction with Epo receptor is discussed. The b inding of Epo to PC12 cells caused a rapid increase in the cytosolic c oncentration of free calcium. The presence of EGTA had no effect on th e Epo binding but completely inhibited the calcium increase, indicatin g that Epo stimulated the calcium influx from outside of the cells. Th e addition of Epo to the culture media of PC12 cells elevated the intr acellular concentrations of monoamines.