The IL-6/IL-12 family cytokines have unique characteristics in that they consist of two distinct subunits forming a heterodimer and each subunit shares with each other. Epstein-Barr virus-induced gene 3 (EBI3) is a common subunit shared by IL-27 and IL-35. Here, we have uncovered a novel role of EBI3 as an intracellular molecule independently of these cytokines. Initially, we found that EBI3 expression is induced in naive CD4+ T cells after stimulation with plate-coated anti-CD3 plus anti-CD28. To investigate the role of EBI3 in naive CD4+ T cells, we then utilized T cell-dependent colitis model, which is induced by transfer of naive CD4+CD25−CD62L+ T cells into RAG2-deficient mice. EBI3-decificent CD4+ T cells failed to induce the colitis with reduced IFN-γ production in CD4+ T cells of intestinal lamina propria. IFN-γ production was also decreased in EBI3-deficient CD4+ T cells differentiated under the pathogenic Th17 polarizing conditions with IL-23 in vitro, whereas this effect was not mediated by a soluble factor. In the EBI3-deficient CD4+ T cells, IL-23 receptor (R) expression was greatly reduced at protein level but not at mRNA level due to increased degradation of IL-23R at proteasome but not at lysosome. Moreover, forced expression of EBI3 in HEK293T cells augmented IL-23R expression via binding to a molecular chaperone, calnexin, and IL-23R. Interestingly, genome-wide association studies and targeted re-sequencing studies recently revealed that the IL-23R variant G149R is linked to protection against the development of inflammatory bowel diseases in humans. Forced expression of EBI3 failed to augment the IL-23R variant G149R expression, due to significantly reduced binding of EBI3 to the variant. Taken together, the present study revealed a novel role for EBI3 as an intracellular molecule that promotes the proper protein folding of IL-23R by binding to calnexin and IL-23R through augmenting its chaperone activity, resulting in development of colitis.