Mycobacterium tuberculosis infection is one of the leading causes of death worldwide. Recognition of pathogen by pattern recognition receptors is crucial for the activation of both innate and adaptive immune responses. Nucleotide-binding oligomerization domain (Nod) 1 and Nod2 are cytoplasmic receptors which detect unique muropeptide from bacterial peptidoglycan. Although Nod2 is critical for initiation of host immune responses against M. tuberculosis infection, the role of Nod1 remains largely unknown. In this study, we investigated the role of Nod1 on cytokines production by bone marrow-derived macrophages (BMDMs) in response to M. tuberculosis. We found that pro-inflammatory cytokines such as IL-6, TNF-α, and IL-1β were induced in BMDMs infected with M. tuberculosis, which was not affected by Nod1 deficiency. Activation of NF-κB and MAPKs by M. tuberculosis was also comparable between WT and Nod1-deficient BMDMs. Interestingly, IL-6 and IL-1β production was reduced in Nod1/Nod2-double deficient BMDMs, as compared with Nod2 single deficient cells. Furthermore, when TLRs signaling was inhibited by LPS pre-treatment, such cytokines production was diminished in Nod1-deficient BMDMs, as compared with WT cells. Our results indicate that Nod1 cooperates with Nod2 or TLRs to produce cytokines in macrophages in response to M. tuberculosis.