A Low-Affinity Human Granulocyte-Macrophage Colony-Stimulating Factor/Murine Erythropoietin Hybrid Receptor Functions in Murine Cell Lines

To identify domains in hematopoietic growth factor receptors that are important for signal transduction, a hybrid receptor (GMER) was constructed by splicing the DNA of the entire extracellular and transmembrane domains of the human granulocyte-macrophage colony-stimulating factor (GM-CSF) receptor...

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Bibliographic Details
Published in:Blood 1993-02, Vol.81 (3), p.587-591
Main Authors: Jubinsky, Paul T., Nathan, David G., Wilson, Donella J., Sieff, Colin A.
Format: Article
Language:English
Subjects:
Animals
Biological and medical sciences
Cell differentiation, maturation, development, hematopoiesis
Cell Division
Cell Line
Cell physiology
Fluorescent Antibody Technique
Fundamental and applied biological sciences. Psychology
Glycophorins - analysis
Glycophorins - biosynthesis
Granulocyte-Macrophage Colony-Stimulating Factor - metabolism
Humans
Interleukin-3 - pharmacology
Macromolecular Substances
Mice
Molecular and cellular biology
Molecular Weight
Receptors, Erythropoietin - genetics
Receptors, Erythropoietin - isolation & purification
Receptors, Erythropoietin - metabolism
Receptors, Granulocyte-Macrophage Colony-Stimulating Factor - genetics
Receptors, Granulocyte-Macrophage Colony-Stimulating Factor - isolation & purification
Receptors, Granulocyte-Macrophage Colony-Stimulating Factor - metabolism
Recombinant Fusion Proteins - isolation & purification
Recombinant Fusion Proteins - metabolism
Transfection
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Summary:To identify domains in hematopoietic growth factor receptors that are important for signal transduction, a hybrid receptor (GMER) was constructed by splicing the DNA of the entire extracellular and transmembrane domains of the human granulocyte-macrophage colony-stimulating factor (GM-CSF) receptor a2 subunit (GMR) to the cytoplasmic domain of the murine erythropoietin receptor (mEpoR). The hybrid receptor was introduced into the interleukin-3 factor-dependent murine hematopoietic cell line Ba/F3. Cells that expressed high receptor numbers were selected by cell sorting using phycoerythrin-labeled human GM-CSF. Immunoprecipitation of GMER from Ba/F3 cells showed a band with an Mr of 105,000 daltons. Human GM-CSF binding to Ba/F3 cells that expressed GMER showed a kd of 3.0 nmol/L and 475 binding sites/cell, while the same cells that expressed GMR had 300 sites/cell and a kd of 3.5 nmol/L. The proliferative response to GM-CSF of Ba/F3 cells that expressed GMER showed ½ maximal cell growth (as measured by 3H-thymidine incorporation) at a GM-CSF concentration of 2.5 × 10–8 mol/L. When cultured in human GM-CSF, Ba/F3-GMER cells expressed cell surface glycophorin. Similar results were obtained with Ba/F3 cells transfected with the mEpoR and cultured in erythropoietin. Expression of GMR plus the human GM-CSF receptor β chain in the same cell line also resulted in human GM-CSF stimulated proliferation; however, cell surface glycophorin was not detected. These data show that a low-affinity GM-CSF/Epo hybrid receptor can promote GM-CSF-dependent proliferation and can induce the expression of glycophorin, an erythroid-specific protein.
ISSN:0006-4971
1528-0020
DOI:10.1182/blood.V81.3.587.587