Cytoplasmic leakage of cathepsins has been associated with the activation of NLRP3 inflammasome-associated caspase-1 [20] . Our proteome analysis showed a rapid increase in the amount of cathepsins in the cytoplasm as a result of virus infection. In addition, secretome analysis showed that there is a major increase in the secretion of cathepsins and their regulators in macrophages infected with influenza A virus. To characterize the role of cathepsins in influenza A virus-induced inflammasome activation we performed Western blot analysis which showed that mature 25 kDa cathepsin B and 34 kDa cathepsin D are secreted simultaneously with biologically active p10 form of caspase-1 in response to influenza A virus infection (Fig. 4A ). Next, macrophages were infected in the presence and absence of cathepsin B inhibitors, Ca-074-Me and z-FA-fmk, after which IL-18 secretion was analysed by ELISA. Both Ca-074-Me (Fig. 4B ) and z-FA-fmk (Fig. S3A ) almost completely abolished IL-18 secretion in response to influenza A virus (Udorn/72) infection. This result was confirmed by Western blot analysis of concentrated cell culture supernatants which showed that the appearance of biologically active form of IL-18, IL-18 p18, is abolished in macrophages that have been infected with influenza A virus in the presence of Ca-074-Me (Fig. 4C ). In accordance with these results, we did not detect any caspase-1 p10 in cell culture supernatants after Ca-074-Me treatment (Fig. 4C ). In addition, Ca-074-Me clearly inhibited IL-18 secretion in response to another influenza A H3N2 strain, Beijing 353/89 (Fig. S3B) . Collectively, our data shows that cathepsin B activity is essential for inflammasome activation in response to influenza A virus infection.