Purification of erythroid progenitor cells and characterization of erythropoietin receptors.
Highly purified murine and human erythroid progenitor cells at the colony-forming unit-erythroid (CFU-E) stage of development were prepared and recombinant human erythropoietin (rEp) was radioiodinated with retention of full biological activity. Specific binding of 125I-rEp to the murine cells revealed 950 receptors on the cell surface. Three hundred had a Kd of 0.09 nM while the remaining receptors had a Kd of 0.57 nM. The human erythroid progenitor cells also had two classes of receptors with a similar number per cell and similar distribution. The high affinity receptors had a Kd of 0.15 nM while the remaining receptors had a Kd of 0.37 nM. 125I-rEp was rapidly internalized into the cells at 37 C and metabolized to iodotyrosine. When cross-linking of 125I-rEp to the murine erythroid progenitor cells was performed with disuccinimidyl suberate, two labelled bands of 100 and 85 kDa were demonstrated. The radioactivity of both bands was reduced when binding was performed in the presence of excessive unlabelled rEp indicating a specific interaction of 125I-rEp with the receptor.
Krantz
S B
SB
Department of Medicine, Vanderbilt University School of Medicine, Nashville, TN.
Sawyer
S T
ST
Sawada
K I
KI
D000740:Anemia / Q000097:blood; D000818:Animals; D002469:Cell Separation / Q000379:methods; D003114:Colony-Forming Units Assay; D004305:Dose-Response Relationship, Drug; D004912:Erythrocytes / Q000166:cytology* / Q000378:metabolism; D004921:Erythropoietin / Q000097:blood*; D005622:Friend murine leukemia virus; D006801:Humans; D007942:Leukemia, Experimental / Q000097:blood; D051379:Mice; D011869:Radioligand Assay; D011956:Receptors, Cell Surface / Q000032:analysis* / Q000378:metabolism; D017467:Receptors, Erythropoietin; D013234:Stem Cells / Q000166:cytology* / Q000378:metabolism; D013997:Time Factors
D011956:Receptors, Cell Surface; D017467:Receptors, Erythropoietin; D004921:Erythropoietin