Interleukin (IL)-17 is a recently described cytokine involved in the amplification of inflammatory responses and pathologies. A hallmark feature of IL-17 is its ability to induce expression of other cytokines and chemokines. In addition, IL-17 potently synergizes with tumor necrosis factor-alpha (TNFalpha) to up-regulate expression of many target genes, particularly IL-6. Despite the many observations of IL-17 signaling synergy observed to date, little is known about the molecular mechanisms that underlie this phenomenon. In the osteoblastic cell line MC-3T3, we have found that IL-17 and TNFalpha exhibit potent synergy in mediating IL-6 secretion. Here, we show that at least part of the functional cooperation between IL-17 and TNFalpha occurs at the level of IL-6 gene transcription. Both the NF-kappaB and CCAAT/enhancer-binding protein (C/EBP; NF-IL6) sites in the IL-6 promoter are important for cooperative gene expression, but NF-kappaB does not appear to be the direct target of the combined signal. Microarray analysis using the Affymetrix mouse MG-U74v2 chip identified C/EBPdelta as another gene target of combined IL-17- and TNFalpha-induced signaling. Because C/EBP family members are known to control IL-6, we examined whether enhanced C/EBPdelta expression is involved in the cooperative up-regulation of IL-6 by IL-17 and TNFalpha. Accordingly, we show that C/EBPdelta (or the related transcription factor C/EBPbeta) is essential for expression of IL-6. Moreover, overexpression of C/EBPdelta (and, to a lesser extent, C/EBPbeta) could substitute for the IL-17 signal at the level of IL-6 transcription. Thus, C/EBP family members, particularly C/EBPdelta, appear to be important for the functional cooperation between IL-17 and TNFalpha.