NLRP3 inflammasome has been shown to be involved in antiviral and antibacterial innate immunity, as well as in the response to adjuvants critical for adaptive immunity. However, inappropriate activation of NLRP3 contributes to deleterious inflammatory syndromes. Although recent studies show that NLRP3 undergoes ubiquitination, the mechanisms for negatively regulating the activation of the NLRP3 inflammasome in response to PAMPs or endogenous DAMPs are largely unknown. In this study, we found that Cbl-b selectively inhibits NLRP3 inflammasome activation, leaving the other key innate response pathways. Further indication of a negative regulation by Cbl-b on NLRP3 inflammasome activity is the finding that LPS injection induces a significant mortality in Cbl-b-/- mice compared to wild-type (WT) controls, and this increased mortality in Cbl-b-/- mice is blocked by an IL-1 receptor antagonist. At the molecular levels, Cbl-b binds to NLRP3 upon LPS priming and ATP stimulation independently of ASC. NLRP3 undergoes ubiquitination in WT macrophages upon NLRP3 inflammasome stimulation, whereas its ubiquitination is impaired in Cbl-b-/- macrophages. Interestingly, NLRP3 inflammasome stimulation also induces Cbl-b tyrosine phosphorylation, which is critical for its ubiquitin ligase activity, thus initiating a negative feedback that controls NLRP3 inflammasome activity. Therefore, we provide a previously-unappreciated function for Cbl-b in regulating NLRP3 inflammasome activation.