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 alpha 2 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 1/2 maximal cell growth (as measured by 3H-thymidine incorporation) at a GM-CSF concentration of 2.5 x 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 beta 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.
Jubinsky
P T
PT
Division of Pediatric Hematology and Oncology, Children's Hospital, Boston, MA.
Nathan
D G
DG
Wilson
D J
DJ
Sieff
C A
CA
D000818:Animals; D002455:Cell Division; D002460:Cell Line; D005455:Fluorescent Antibody Technique; D006021:Glycophorins / Q000032:analysis / Q000096:biosynthesis; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000378:metabolism; D006801:Humans; D007377:Interleukin-3 / Q000494:pharmacology; D046911:Macromolecular Substances; D051379:Mice; D008970:Molecular Weight; D017467:Receptors, Erythropoietin / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D016187:Receptors, Granulocyte-Macrophage Colony-Stimulating Factor / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000302:isolation & purification / Q000378:metabolism; D014162:Transfection
D006021:Glycophorins; D007377:Interleukin-3; D046911:Macromolecular Substances; D017467:Receptors, Erythropoietin; D016187:Receptors, Granulocyte-Macrophage Colony-Stimulating Factor; D011993:Recombinant Fusion Proteins; D016178:Granulocyte-Macrophage Colony-Stimulating Factor