title: Dynamics of the gp130 cytokine complex: a model for assembly on the cellular membrane.
abstract: Cytokines of the interleukin-6 (IL-6)-type family all bind to the glycoprotein gp130 on the cell surface and require interaction with two gp130 or one gp130 and another related signal transducing receptor subunit. In addition, some cytokines of this family, such as IL-6, interleukin-11, ciliary neurotrophic factor, neuropoietin, cardiotrophin-1, and cardiotrophin-1-like-cytokine, interact with specific ligand binding receptor proteins. High- and low-affinity binding sites have been determined for these cytokines. So far, however, the stoichiometry of the signaling receptor complexes has remained unclear, because the formation of the cytokine/cytokine-receptor complexes has been analyzed with soluble receptor components in solution, which do not necessarily reflect the situation on the cellular membrane. Consequently, the binding affinities measured in solution have been orders of magnitude below the values obtained with whole cells. We have expressed two gp130 extracellular domains in the context of a Fc-fusion protein, which fixes the receptors within one dimension and thereby restricts the flexibility of the proteins in a fashion similar to that within the plasma membrane. We measured binding of IL-6 and interleukin-b receptor (IL-6R) by means of fluorescence-correlation spectroscopy. For the first time we have succeeded in recapitulating in a cell-free condition the binding affinities and dynamics of IL-6 and IL-6R to the gp130 receptor proteins, which have been determined on whole cells. Our results demonstrate that a dimer of gp130 first binds one IL-6/IL-6R complex and only at higher ligand concentrations does it bind a second IL-6/IL-6R complex. This view contrasts with the current perception of IL-6 receptor activation and reveals an alternative receptor activation mechanism.
authors.affiliation: Evotec Technologies GmbH, D-40225 Düsseldorf, Germany.
authors.forename: Andreas
authors.initials: A
authors.lastname: Schroers
authors.forename: Oliver
authors.initials: O
authors.lastname: Hecht
authors.forename: Karl-Josef
authors.initials: KJ
authors.lastname: Kallen
authors.forename: Michael
authors.initials: M
authors.lastname: Pachta
authors.forename: Stefan
authors.initials: S
authors.lastname: Rose-John
authors.forename: Joachim
authors.initials: J
authors.lastname: Grötzinger
mesh_terms: D000818:Animals; D015703:Antigens, CD / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D016466:CHO Cells; D002462:Cell Membrane / Q000378:metabolism*; D002850:Chromatography, Gel; D006224:Cricetinae; D003412:Cricetulus; D050822:Cytokine Receptor gp130; D003627:Data Interpretation, Statistical; D007141:Immunoglobulin Fc Fragments / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D015850:Interleukin-6 / Q000378:metabolism*; D008562:Membrane Glycoproteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D008954:Models, Biological; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D013050:Spectrometry, Fluorescence
chemical_list: D015703:Antigens, CD; D007141:Immunoglobulin Fc Fragments; D015850:Interleukin-6; D008562:Membrane Glycoproteins; D011993:Recombinant Fusion Proteins; D050822:Cytokine Receptor gp130
references.citation: Biophys J. 2000 Dec;79(6):2858-66
references.pmid: 11106594
references.citation: Stem Cells. 2002;20(2):119-38
references.pmid: 11897869
references.citation: J Biol Chem. 1999 Apr 23;274(17):11859-67
references.pmid: 10207005
references.citation: J Mol Biol. 1994 Sep 30;242(4):497-507
references.pmid: 7932706
references.citation: Blood. 2003 Aug 15;102(4):1186-95
references.pmid: 12714508
references.citation: Science. 2003 Jun 27;300(5628):2101-4
references.pmid: 12829785
references.citation: Biol Chem. 1999 Jul-Aug;380(7-8):803-13
references.pmid: 10494829
references.citation: J Exp Med. 1996 Apr 1;183(4):1399-406
references.pmid: 8666898
references.citation: Trends Biotechnol. 2002 Oct;20(10):417-9
references.pmid: 12220903
references.citation: Eur Cytokine Netw. 2000 Mar;11(1):27-38
references.pmid: 10705296
references.citation: Mol Cell. 2003 Sep;12(3):577-89
references.pmid: 14527405
references.citation: Neuron. 1996 Jul;17(1):63-74
references.pmid: 8755479
references.citation: Biophys J. 1976 Nov;16(11):1315-29
references.pmid: 974223
references.citation: Blood. 1998 Nov 15;92(10):3495-504
references.pmid: 9808540
references.citation: J Exp Med. 1997 Feb 17;185(4):755-66
references.pmid: 9034153
references.citation: Eur J Biochem. 2001 Jan;268(1):160-7
references.pmid: 11121117
references.citation: EMBO J. 1995 May 1;14(9):1942-51
references.pmid: 7744001
references.citation: Mol Cell Proteomics. 2002 Oct;1(10):805-15
references.pmid: 12438563
references.citation: Annu Rev Immunol. 1997;15:797-819
references.pmid: 9143707
references.citation: Eur J Neurosci. 1999 Sep;11(9):2995-3004
references.pmid: 10510164
references.citation: Nat Cell Biol. 2000 Mar;2(3):168-72
references.pmid: 10707088
references.citation: Science. 2001 Mar 16;291(5511):2150-5
references.pmid: 11251120
references.citation: J Biol Chem. 1996 Dec 20;271(51):32635-43
references.pmid: 8999038
references.citation: EMBO J. 1996 Jun 3;15(11):2726-37
references.pmid: 8654370
references.citation: J Biol Chem. 1997 Jun 20;272(25):15760-4
references.pmid: 9188471
references.citation: Proc Natl Acad Sci U S A. 2004 Apr 6;101(14):4827-32
references.pmid: 15051883
references.citation: Proc Natl Acad Sci U S A. 1998 Mar 17;95(6):3251-6
references.pmid: 9501249
references.citation: J Biol Chem. 1994 Sep 16;269(37):23286-9
references.pmid: 8083235
references.citation: J Immunol. 1992 Sep 15;149(6):2021-7
references.pmid: 1381393
references.citation: J Immunol. 1999 Oct 15;163(8):4583-9
references.pmid: 10510402
references.citation: Science. 1988 Aug 12;241(4867):825-8
references.pmid: 3136546
references.citation: FEBS Lett. 1992 Jul 20;306(2-3):219-22
references.pmid: 1321736
references.citation: Science. 1999 Feb 12;283(5404):987-90
references.pmid: 9974392
references.citation: Cell. 1989 Aug 11;58(3):573-81
references.pmid: 2788034
references.citation: Biochem J. 1994 Jun 1;300 ( Pt 2):281-90
references.pmid: 8002928
references.citation: J Biomol Screen. 2001 Feb;6(1):29-37
references.pmid: 11679163
references.citation: J Immunol. 2000 Dec 15;165(12):7042-9
references.pmid: 11120832
references.citation: Adv Exp Med Biol. 2001;495:145-51
references.pmid: 11774558
references.citation: Biochim Biophys Acta. 2002 Nov 11;1592(3):251-63
references.pmid: 12421670
references.citation: Nat Biotechnol. 1997 Feb;15(2):142-5
references.pmid: 9035138
references.citation: Anal Biochem. 1993 May 1;210(2):425-8
references.pmid: 8512082
references.citation: Drug Discov Today. 2001 Jun 1;6(12):637-646
references.pmid: 11408200
references.citation: J Immunol. 1999 Feb 1;162(3):1480-7
references.pmid: 9973404
references.citation: Cytokine. 1992 Mar;4(2):96-100
references.pmid: 1633265
references.citation: Eur J Biochem. 1995 Dec 1;234(2):661-9
references.pmid: 8536717
references.citation: J Biol Chem. 1996 Aug 16;271(33):20138-44
references.pmid: 8702737