Importance of the membrane-proximal extracellular domains for activation of the signal transducer glycoprotein 130

The transmembrane glycoprotein gp130 is the common signal transducing recep tor subunit of the IL-6-type cytokines, The gp130 extracellular part is pre dicted to consist of six individual domains. Whereas the role of the three membrane-distal domains (D1-D3) in binding of IL-6 and IL-11 is well establ ished, the function of the membrane-proximal domains (D4-D6) is unclear. Ma pping of a neutralizing mAb to the membrane-proximal part of gp130 suggests a functional role of D4-D6 in receptor activation. Individual deletion of these three domains differentially interferes with ligand binding of the so luble and membrane-bound receptors, All deletion mutants do not signal in r esponse to IL-6 and IL-11. The deletion mutants Delta 4 and, to a lesser ex tent, Delta 6 are still activated by agonistic monoclonal gp130 Abs, wherea s the deletion mutant Delta 5 does not respond. Because membrane-bound Delt a 5 binds IL-6/soluble IL-6R as does wild-type gp130, but does not transduc e a signal in response to various stimuli, this domain plays a prominent ro le in coupling of ligand binding and signal transduction. Replacement of th e fifth domain of gp130 by the corresponding domain of the homologous G-CSF receptor leads to constitutive activation of the chimera upon overexpressi on in COS-7 cells. In HepG2 cells this mutant responds to IL-6 comparable t o wild-type gp130, Our findings suggest a functional role of the membrane-p roximal domains of gp130 in receptor activation. Thus, within the hematopoi etic receptor family the mechanism of receptor activation critically depend s on the architecture of the receptor ectodomain.