Granulocyte-macrophage colony stimulating factor receptor alpha and beta chain complexes can form both high and intermediate affinity functional receptors.
Scatchard analysis of primary human haemopoietic cells using iodinated GM-CSF suggests that there are low, intermediate and high affinity classes of the GM-CSF receptor. To investigate the molecular basis of this, we generated a clone of transfected NIH3T3 cells that constitutively expressed the human granulocyte-macrophage colony stimulating factor receptor (GM-CSF R) beta chain and inducibly expressed the human GM-CSF R alpha chain. In the cells fully induced to express the alpha chain the overall level of expression of the alpha and beta chains at the cell surface was comparable with that found in primary haemopoietic cells and cell lines. When cells were partially induced to express the alpha chain, the alpha:beta ratio determined by antibody binding was approximately 1:1 and Scatchard analysis revealed a single class of intermediate affinity receptors (Kd = 614+/-88 pM). In cells with fully induced alpha chain expression, the alpha:beta ratio was approximately 3:1 and there was a switch to a dual high and low affinity receptor with K(d)s of 67+/-32 pM and 1.7+/-0.56 nM respectively. The change from intermediate to high affinity was not associated with changes in alphabeta stoichiometry as detected by cross-linking with radiolabelled GM-CSF and gel electrophoresis. Both the high and intermediate affinity receptors were able to activate the STAT 5 and the MAP kinase pathways, although there was a difference in the ligand dose-response curves which was compatible with the different affinities of the receptors. It is proposed that the switch from an intermediate to high affinity receptor was due to the availability of free alpha chains presenting ligand to the alphabeta chain complexes at the surface of the cell membrane.
Wheadon
H
H
Department of Haematology, University College London Medical School.
Devereux
S
S
Khwaja
A
A
Linch
D C
DC
D016475:3T3 Cells; D000818:Animals; D015153:Blotting, Western; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism; D006801:Humans; D051379:Mice; D008894:Milk Proteins; D010766:Phosphorylation; D016187:Receptors, Granulocyte-Macrophage Colony-Stimulating Factor / Q000378:metabolism*; D050799:STAT5 Transcription Factor; D015398:Signal Transduction; D015534:Trans-Activators / Q000378:metabolism
D004268:DNA-Binding Proteins; D008894:Milk Proteins; D016187:Receptors, Granulocyte-Macrophage Colony-Stimulating Factor; D050799:STAT5 Transcription Factor; D015534:Trans-Activators; D017871:Calcium-Calmodulin-Dependent Protein Kinases