The hepatic interleukin-6 receptor. Studies on its structure and regulation by phorbol 12-myristate 13-acetate-dexamethasone.
Recombinant human 125I-interleukin-6 (IL-6) was cross-linked with the homobifunctional reagent disuccinimidyl suberate to human hepatoma cells (HepG2). Three recombinant human 125I-IL-6-containing complexes of apparent molecular masses of 100, 120, and 200 kDa were immunoprecipitated with specific antibodies to human IL-6 or to the 80-kDa IL-6 receptor subunit. We show by immunoprecipitation, peptide mapping, and by the use of a cleavable heterobifunctional cross-linker (Denny-Jaffe reagent) that different polypeptides are involved in the formation of the 100- and 120-kDa IL-6-containing complexes. The molecular compositions of the 100- and 120-kDa cross-linked complexes were identified. The 100-kDa complex consisted of one ligand and one IL-6 receptor subunit, glycoprotein 80 (gp80), whereas the 120-kDa complex was found to be composed of one ligand and a polypeptide which was immunoprecipitable with the monoclonal antibody AM64 directed against gp130. Exposure of HepG2 cells to phorbol 12-myristate 13-acetate (PMA) or PMA-dexamethasone led to an increase in the 80-kDa IL-6 receptor mRNA and functional receptor protein. Whereas treatment of HepG2 cells with PMA led to an increase in the formation of gp80.gp130.IL-6 complexes determined by cross-linking, no corresponding increase in high affinity binding sites was found. The existence of a third IL-6 receptor subunit present in limiting amounts on HepG2 cells is proposed to explain this discrepancy. Evidence is presented that the 80-kDa IL-6 receptor up-regulation by PMA-dexamethasone is caused by the depletion of protein kinase C since the protein kinase C inhibitor staurosporine mimics the effect of PMA-dexamethasone.
Pietzko
D
D
Institut für Biochemie, Rheinisch-Westfälische Technische Hochschule Aachen, Federal Republic of Germany.
Zohlnhöfer
D
D
Graeve
L
L
Fleischer
D
D
Stoyan
T
T
Schooltink
H
H
Rose-John
S
S
Heinrich
P C
PC
D002478:Cells, Cultured; D003432:Cross-Linking Reagents; D003907:Dexamethasone / Q000494:pharmacology*; D015536:Down-Regulation; D015850:Interleukin-6 / Q000378:metabolism*; D008099:Liver / Q000166:cytology / Q000378:metabolism*; D011493:Protein Kinase C / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D011971:Receptors, Immunologic / Q000737:chemistry / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D019947:Receptors, Interleukin-6; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*
D003432:Cross-Linking Reagents; D015850:Interleukin-6; D012333:RNA, Messenger; D011971:Receptors, Immunologic; D019947:Receptors, Interleukin-6; D003907:Dexamethasone; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate