NLRs are cytoplasmic proteins that sense cellular stress and intracellular damage resulting from pathogen uptake. To date, the role of NLRs has been studied using combinations of NLR and TLR agonists, but the interplay between two different NLRs remains uncharacterized. In this study, we employed microarrays to investigate in DCs the regulation of gene transcription mediated by activation of NOD2 and NLRP3 pathways using MDP and MSU. MDP and MSU co-stimulation of murine BMDCs up-regulated the expression of genes encoding molecules for antigen presentation and co-stimulation (MHC class II, CD80, CD86), integrins (ITGB3, ITGAV), cytokines (IL-1α, IL-1β, IL-6, IL-2, IL-23p19, IL-12p40), and chemokines (CXCL1, CXCL2). Transcription of the cytokine genes induced by MDP and MSU partially depended on NOD2 but was independent of NLRP3. Finally, we showed that ERK1 and c-JUN activation increased upon MDP and MSU co-stimulation. As a whole, the results indicate that two different NLR activators synergize at the transcriptional level, leading to unique differential expression of genes involved in the innate immune response.