Autophagy is a key regulator of cellular homeostasis that can be activated by pathogen-associated molecules and has recently been shown to influence IL-1b secretion by macrophages. However, the mechanisms behind this are unclear. Here, we describe a novel role for autophagy in regulating the production of IL-1b in antigen-presenting cells. After treatment of macrophages with Toll-like receptor (TLR) ligands, pro-IL-1b was specifically sequestered into autophagosomes, while further activation of autophagy with rapamycin induced the degradation of pro-IL-1b and blocked secretion of the mature cytokine. Inhibition of autophagy promoted the processing and secretion of IL-1b by antigen-presenting cells in a NLRP3- and TRIF-dependent manner. This effect was reduced by inhibition of reactive oxygen species (ROS), but was independent of NOX2. Induction of autophagy in mice in vivo with rapamycin reduced serum levels of IL-1b in response to challenge with LPS. These data demonstrate that autophagy controls the production of IL-1b through at least two separate mechanisms; by targeting pro-IL-1 for lysosomal degradation and by regulating activation of the NLRP3 inflammasome.