Characterization of three erythropoietin (Epo)-binding proteins in various human Epo-responsive cell lines and in cells transfected with human Epo-receptor cDNA.
Molecular cloning of a cDNA for a mouse erythropoietin (Epo) receptor (EpoR) has facilitated the understanding of the structure of this receptor. However, there is, as yet, no explanation for the discrepancy between the protein recognized by specific antibodies against mouse EpoR and the unexpectedly larger species that can be cross-linked to labeled Epo. It is unclear whether the product of an unidentified gene is included in the EpoR complex. In the present study, we directly compared the cross-linking patterns for human EpoR that were endogenously expressed in three types of Epo-responsive cell, and that was artificially expressed in nonhematopoietic cells after transfection with cDNA for human EpoR. We observed that 85-kD and 105-kD proteins formed ligand-receptor complexes in all the human Epo-responsive cells and, furthermore, that the formation of a complex derived from the 70-kD protein was dependent on the level of expression of the cloned EpoR mRNA in these cells. By contrast, a prominent cross-linked band derived from the 70-kD protein and a weaker band derived from the 80- to 85-kD protein, but no band derived from the 105-kD protein, could be shown in the case of nonhematopoietic cells transfected with the human EpoR cDNA. These observations suggest that the cloned cDNA for human EpoR alone does not allow generation of the complete EpoR in nonhematopoietic cells and that the 105-kD Epo-binding protein may represent the product of an as yet unidentified gene that is expressed in hematopoietic cells.
Takahashi
T
T
Third Department of Internal Medicine, Faculty of Medicine, University of Tokyo, Japan.
Chiba
S
S
Hirano
N
N
Yazaki
Y
Y
Hirai
H
H
D000650:Amnion; D000818:Animals; D015152:Blotting, Northern; D015139:Blotting, Southern; D002460:Cell Line; D002522:Chlorocebus aethiops; D003432:Cross-Linking Reagents; D018076:DNA, Complementary / Q000235:genetics*; D004921:Erythropoietin / Q000378:metabolism / Q000494:pharmacology*; D015870:Gene Expression; D006801:Humans; D007163:Immunosorbent Techniques; D007457:Iodine Radioisotopes; D007668:Kidney; D004915:Leukemia, Erythroblastic, Acute; D051379:Mice; D008970:Molecular Weight; D012333:RNA, Messenger / Q000032:analysis; D017467:Receptors, Erythropoietin / Q000737:chemistry / Q000235:genetics*; D014162:Transfection; D014407:Tumor Cells, Cultured
D003432:Cross-Linking Reagents; D018076:DNA, Complementary; D007457:Iodine Radioisotopes; D012333:RNA, Messenger; D017467:Receptors, Erythropoietin; D004921:Erythropoietin