The soluble interleukin-6 receptor is generated by shedding.
The ligand-binding subunit (gp80) of the human interleukin-6 receptor (IL-6R) was transiently expressed in COS-7 cells. The metabolically labeled protein was shown to be quantitatively released from the membrane within 20 h. We identified the protein released from the transfected COS-7 cells after purification to homogeneity and N-terminal sequencing as a soluble form of the gp80/IL-6R. Shedding of the gp80 protein was strongly induced by 4 beta-phorbol-12-myristate-13-acetate, indicating that the process was regulated by protein kinase C (PKC). This was further corroborated by the finding that co-transfection of a PKC expression plasmid led to enhanced shedding of the gp80 protein. Since shedding of gp80 could not be prevented by treatment of the cells with inhibitors of all known classes of proteases, a novel protease seems to be involved. As a control, an unrelated membrane protein (vesicular stomatitis virus glycoprotein) was transfected into COS-7 cells and analyzed for shedding. Since the turnover of this protein was not mediated by shedding, we conclude that the release of gp80 from COS-7 cells is a specific process. The shed gp80 protein specifically binds IL-6, and this complex shows biological activity on human hepatoma cells. Human peripheral blood monocytes released a soluble form of the gp80 protein into the culture medium upon PMA treatment indicating that PKC-regulated shedding is the physiological mechanism of generation of the soluble IL-6R.
Müllberg
J
J
Institut für Biochemie, RWTH Aachen, FRG.
Schooltink
H
H
Stoyan
T
T
Günther
M
M
Graeve
L
L
Buse
G
G
Mackiewicz
A
A
Heinrich
P C
PC
Rose-John
S
S
D002460:Cell Line; D002478:Cells, Cultured; D004591:Electrophoresis, Polyacrylamide Gel; D005455:Fluorescent Antibody Technique; D015870:Gene Expression; D006801:Humans; D015850:Interleukin-6 / Q000378:metabolism; D008562:Membrane Glycoproteins / Q000302:isolation & purification / Q000378:metabolism; D009000:Monocytes / Q000187:drug effects / Q000378:metabolism; D010957:Plasmids; D011480:Protease Inhibitors / Q000494:pharmacology; D011493:Protein Kinase C / Q000378:metabolism; D011971:Receptors, Immunologic / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D019947:Receptors, Interleukin-6; D012995:Solubility; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014162:Transfection; D014759:Viral Envelope Proteins / Q000378:metabolism
C028391:G protein, vesicular stomatitis virus; C508603:IL6R protein, human; D015850:Interleukin-6; D008562:Membrane Glycoproteins; D011480:Protease Inhibitors; D011971:Receptors, Immunologic; D019947:Receptors, Interleukin-6; D014759:Viral Envelope Proteins; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate