Molecular characterization of grass carp interleukin-6 receptor and the agonistic activity of its soluble form in head kidney leucocytes

Abstract

Interleukin (IL)-6 receptor (IL-6R) can specifically bind to IL-6 and the complex subsequently recruits a transmembrane signal transducer, gp130, to trigger the intracellular signal transduction. IL-6R exists in two forms, a transmembrane IL-6R and a soluble IL-6R (sIL-6R), leading to different signal transduction mechanisms as classic signaling and trans-signaling, respectively. There is now a general consensus that these two modes of signal transduction can mediate anti-inflammatory and pro-inflammatory activities of IL-6. The study on Il-6r is limited although Il-6 has been well studied in teleost. In the present study, a cDNA encoding grass carp Il-6r (gcIl-6r) was isolated. An in-silico analysis showed that gcIl-6r shared the same functional domains and conserved gene synteny at its loci with mouse homologue, and its amino acid sequence was conserved in fish species. A tissue distribution assay demonstrated that gcil6r mRNA was expressed with high levels in immune tissues including spleen and head kidney, and its expression was induced by LPS and Poly I:C in grass carp head kidney leucocytes (HKLs). An in vitro binding assay showed that recombinant soluble gcIl-6r (rgcsIl-6r) could specifically bind to recombinant gcIl-6 (rgcIl-6) protein. Moreover, rgcIl-6 stimulated suppressor of cytokine signaling 3 (socs3)'s mRNA expression in grass carp HKLs and it combined with rgcsIl-6r increased socs3 mRNA expression in CIK cells with gp130 but without Il-6r expression. In HKLs, rgcIl-6 stimulated the mRNA levels of both pro-inflammatory (tnfa and il1b) and anti-inflammatory (il10) cytokines, and rgcsIl-6r could augment these stimulatory effects of gcIl-6. Taken these data together, gcsIl-6r can mediate the immuno-regulatory functions of gcIl-6 and has an agonistic property in these actions of Il-6 in grass carp.