Granulocyte-macrophage colony-stimulating factor (GM-CSF) and inflammatory stimuli up-regulate secretion of the soluble GM-CSF receptor in human monocytes: evidence for ectodomain shedding of the cell surface GM-CSF receptor alpha subunit.
Soluble GM-CSF receptor alpha subunit (sGMRalpha) is a soluble isoform of the GMRalpha that is believed to arise exclusively through alternative splicing of the GMRalpha gene product. The sGMRalpha mRNA is expressed in a variety of tissues, but it is not clear which cells are capable of secreting the protein. We show here that normal human monocytes, but not lymphocytes, constitutively secrete sGMRalpha. Stimulation of monocytes with GM-CSF, LPS, PMA, or A23187 rapidly up-regulates the secretion of sGMRalpha in a dose-dependent manner, demonstrating that secretion is also regulated. To determine whether sGMRalpha arose exclusively through alternative splicing of the GMRalpha gene product, or whether it could also be generated through ectodomain shedding of GMRalpha, we engineered a murine pro-B cell line (Ba/F3) to express exclusively the cDNA for cell surface GMRalpha (Ba/F3.GMRalpha). The Ba/F3.GMRalpha cell line, but not the parental Ba/F3 cell line, constitutively shed a sGMRalpha-like protein that bound specifically to GM-CSF, was equivalent in size to recombinant alternatively spliced sGMRalpha (60 kDa), and was recognized specifically by a mAb raised against the ectodomain of GMRalpha. Furthermore, a broad-spectrum metalloprotease inhibitor (BB94) reduced constitutive and PMA-, A23187-, and LPS-induced secretion of sGMRalpha by monocytes, suggesting that shedding of GMRalpha by monocytes may be mediated in part through the activity of metalloproteases. Taken together, these observations demonstrate that sGMRalpha is constitutively secreted by monocytes, that GM-CSF and inflammatory mediators up-regulate sGMRalpha secretion, and that sGMRalpha arises not only through alternative splicing but also through ectodomain shedding of cell surface GMRalpha.
Prevost
Jay M
JM
Cancer Biology Research Group, Southern Alberta Cancer Research Center, University of Calgary, 3330 Hospital Drive NW, Calgary, Alberta, Canada T2N 4N1.
Pelley
Jennifer L
JL
Zhu
Weibin
W
D'Egidio
Gianni E
GE
Beaudry
Paul P
PP
Pihl
Carin
C
Neely
Graham G
GG
Claret
Emmanuel
E
Wijdenes
John
J
Brown
Christopher B
CB
D017398:Alternative Splicing / Q000276:immunology; D000818:Animals; D000001:Calcimycin / Q000494:pharmacology; D002462:Cell Membrane / Q000276:immunology / Q000378:metabolism; D002478:Cells, Cultured; D006224:Cricetinae; D010450:Endopeptidases / Q000378:metabolism; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000378:metabolism / Q000494:pharmacology*; D006801:Humans; D006868:Hydrolysis; D018836:Inflammation Mediators / Q000494:pharmacology*; D008070:Lipopolysaccharides / Q000494:pharmacology; D008214:Lymphocytes / Q000378:metabolism; D051379:Mice; D009000:Monocytes / Q000187:drug effects / Q000276:immunology / Q000378:metabolism* / Q000473:pathology*; D017434:Protein Structure, Tertiary; D021122:Protein Subunits; D016187:Receptors, Granulocyte-Macrophage Colony-Stimulating Factor / Q000037:antagonists & inhibitors / Q000096:biosynthesis / Q000378:metabolism*; D012995:Solubility; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D015854:Up-Regulation / Q000276:immunology*
D018836:Inflammation Mediators; D008070:Lipopolysaccharides; D021122:Protein Subunits; D016187:Receptors, Granulocyte-Macrophage Colony-Stimulating Factor; D000001:Calcimycin; D016178:Granulocyte-Macrophage Colony-Stimulating Factor; D010450:Endopeptidases; D013755:Tetradecanoylphorbol Acetate