Abstract |
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.
|
Authors | D Pietzko, D Zohlnhöfer, L Graeve, D Fleischer, T Stoyan, H Schooltink, S Rose-John, P C Heinrich |
Journal | The Journal of biological chemistry
(J Biol Chem)
Vol. 268
Issue 6
Pg. 4250-8
(Feb 25 1993)
ISSN: 0021-9258 [Print] United States |
PMID | 8440709
(Publication Type: Journal Article, Research Support, Non-U.S. Gov't)
|
Chemical References |
- Cross-Linking Reagents
- Interleukin-6
- RNA, Messenger
- Receptors, Immunologic
- Receptors, Interleukin-6
- Dexamethasone
- Protein Kinase C
- Tetradecanoylphorbol Acetate
|
Topics |
- Cells, Cultured
- Cross-Linking Reagents
- Dexamethasone
(pharmacology)
- Down-Regulation
- Interleukin-6
(metabolism)
- Liver
(cytology, metabolism)
- Protein Kinase C
(metabolism)
- RNA, Messenger
(metabolism)
- Receptors, Immunologic
(chemistry, drug effects, genetics, metabolism)
- Receptors, Interleukin-6
- Tetradecanoylphorbol Acetate
(pharmacology)
|