Lack of the IL-1 receptor accessory protein (IL-1RAcP) abrogates responses to IL-33 and IL-1 in the mouse thymoma clone EL-4 D6/76 cells. Reconstitution with full-length IL-1RAcP is sufficient to restore responsiveness to IL-33 and IL-1. IL-33 activates IL-1 receptor-associated kinase-1, cJun-N-terminal kinase, and the NF-kappaB pathway in an IL-1RAcP-dependent manner and results in IL-2 release. IL-33 is able to induce the release of proinflammatory cytokines in bone marrow-derived (BMD) mast cells, indicating that IL-33 may have a proinflammatory potential like its relatives IL-1 and IL-18, in addition to its Th2-skewing properties in the adaptive response described previously. Blocking of murine IL-1RAcP with the neutralizing antibody 4C5 inhibits response of mouse thymoma cells and BMD mast cells to IL-33. The interaction of either membrane-bound or soluble forms of IL-1RAcP and IL-33Ralpha-chain depends on the presence of IL-33, as demonstrated by coimmunoprecipitation assays. These data demonstrate that IL-1RAcP is indispensable for IL-33 signaling. Furthermore, they suggest that IL-1RAcP is used by more than one alpha-chain of the IL-1 receptor family and thus may resemble a common beta-chain of that family.