A LOW-AFFINITY CHIMERIC HUMAN ALPHA BETA-GRANULOCYTE-MACROPHAGE COLONY-STIMULATING FACTOR-RECEPTOR INDUCES LIGAND-DEPENDENT PROLIFERATION IN A MURINE CELL-LINE/

The high affinity receptor for granulocyte-macrophage colony-stimulati ng factor (GM-CSF) is composed of at least two subunits, an 85-kDa low affinity GM-CSF-binding protein (alpha-GMR) and a 120-kDa beta-subuni t (beta-GMR) necessary for high affinity binding and signal transducti on. Previous studies have shown that deletion of the intracellular do main of alpha-GMR inactivates the receptor's ability to support prolif eration, but has no effect on GM-CSF binding. Using anti-alpha-GMR and anti-beta-GMR-specific antibodies, we show that alpha-GMR and beta-GM R coprecipitate only after GM-CSF binding, suggesting that binding of GM-CSF induces stabilization or assembly of an activated receptor comp lex involving recruitment of beta-GMR chains. To understand the contri bution of each subunit of this receptor to the generation of an activa ted receptor complex, we attempted to construct minimal receptors with some or all of the functions of the wild-type heterodimer. We found t hat a hybrid human alpha/beta-GMR molecule in which the extracellular and transmembrane segments are composed of alpha-GMR sequences and the intracellular segment is composed of beta-GMR bound GM-CSF with low a ffinity, but activated tyrosine kinase activity, induced receptor inte rnalization, and supported short- and long-term proliferation of trans fected Ba/F3 cells. At least 1 ng/ml human GM-CSF was required for gro wth stimulation, and maximal proliferation occurred at a concentration of 10 ng/ml. This was 10-100-fold more than needed to stimulate growt h of Ba/F3 cells expressing both full-length human alpha-GMR and beta- GMR and 1000-fold less than needed to stimulate growth of Ba/F3 cells expressing only human alpha-GMR. These results indicate that the cytop lasmic domain of alpha-GMR is not required to initiate a unique signal ing event for proliferation in Ba/F3 cells, but can be functionally re placed by the cytoplasmic domain of beta-GMR.