Synthetic trimeric interleukin-6 receptor complexes with a STAT3 phosphorylation dominated activation profile.

In Interleukin (IL)-6 signalling, IL-6 site I binds to the IL-6 receptor (IL-6R) first, following by IL-6 site II interaction to domain 2/3 of gp130 to form premature trimeric IL-6:IL-6R:gp130 receptor complexes. Formation of the mature hexameric receptor complex is then facilitated by the inter-trimeric interaction of IL-6 site III with domain 1 of the opposing gp130. The two gp130-associated Janus kinases (JAKs) trans-phosphorylate when their spatiotemporal pairing is correct, which causes the activation of STAT, ERK, and AKT pathways in a balanced manner. Since the intracellular domain (ICD) of IL-6R is not needed for STAT/ERK/AKT phosphorylation, we investigated the conditions under which a chimeric IL-6R-gp130 receptor protein confers biological activity. For IL-6R-gp130, the extracellular domain (ECD) of IL-6R was fused to the transmembrane domain (TMD) and ICD of gp130. Co-expression of IL-6R-gp130 with signalling-deficient gp130 variants did not induce IL-6 signalling, suggesting that the assembly of hexameric complexes failed to dimerize the IL-6R-associated JAKs correctly. By mimicking the premature trimeric receptor complex, IL-6-mediated dimerization of IL-6R-gp130 with the single-cytokine-binding variant gp130 induced signalling. Of note, IL-6 signalling via these synthetic gp130:IL-6R-gp130 complexes resulted predominantly in STAT3 phosphorylation. A STAT3-dominated profile was also observed after IL-6-induced signalling mediated by a JAK-deficient IL-6R-gp130 variant in complex with the JAK-proficient but STAT/ERK/AKT-deficient gp130 variant. Our data showed that effective ERK/AKT signalling could not be executed after intracellular domain swapping from gp130 to the IL-6R. Taken together, the chimeric IL-6R/gp130 receptor may be helpful in the creation of customized synthetic IL-6 signalling.