Identification of the receptor for erythropoietin on human and murine erythroleukemia cells and modulation by phorbol ester and dimethyl sulfoxide.
Erythropoietin, a glycoprotein that regulates erythropoiesis, initiates its biological effects by binding to a cell-surface receptor. Little is known about the structure of the erythropoietin receptor and the events that follow binding of erythropoietin to its receptor, in part because of the difficulty of obtaining sufficient quantities of cells that express the erythropoietin receptor. We used both iodinated and metabolically labeled erythropoietin to characterize the receptor on a variety of erythroleukemia cell lines not previously tested, and we have identified both human and murine cell lines that display large numbers of erythropoietin receptors. Both erythropoietin-responsive and -nonresponsive cell lines exhibit a single class of binding sites. The human erythroleukemia cell line OCIM1 exhibits approximately 3000 erythropoietin receptors per cell with a Kd of 280 pM. The erythropoietin-responsive Rauscher red 5-1.5 murine erythroleukemia cell line displays approximately 1700 receptors per cell with a Kd of 440 pM. The GM979 murine erythroleukemia cell line has approximately 1600 receptors per cell with a Kd of 660 pM. Induction of the erythroid phenotype by dimethyl sulfoxide or its suppression by phorbol 12-myristate 13-acetate was accompanied by an increase or decrease, respectively, in erythropoietin receptor number. Affinity crosslinking of labeled erythropoietin to the receptor identified two proteins corresponding to estimated molecular masses of 95 and 105 kDa. The OCIM1, Rauscher, and GM979 erythroleukemia cell lines provide a useful model for the study of postreceptor signaling events, as well as a convenient source for purification of the erythropoietin receptor.
Broudy
V C
VC
Division of Hematology and Metabolism, University of Washington, Seattle 98195.
Lin
N
N
Egrie
J
J
de Haën
C
C
Weiss
T
T
Papayannopoulou
T
T
Adamson
J W
JW
D000818:Animals; D002460:Cell Line; D004121:Dimethyl Sulfoxide / Q000494:pharmacology*; D004921:Erythropoietin / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D004915:Leukemia, Erythroblastic, Acute / Q000378:metabolism*; D051379:Mice; D008970:Molecular Weight; D011956:Receptors, Cell Surface / Q000187:drug effects / Q000302:isolation & purification / Q000378:metabolism*; D017467:Receptors, Erythropoietin; D011994:Recombinant Proteins / Q000378:metabolism*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*
D011956:Receptors, Cell Surface; D017467:Receptors, Erythropoietin; D011994:Recombinant Proteins; D004921:Erythropoietin; D013755:Tetradecanoylphorbol Acetate; D004121:Dimethyl Sulfoxide