Mycobacterial EST12 activates a RACK1–NLRP3–gasdermin D pyroptosis–IL-1β immune pathway

Abstract

Pyroptosis, an inflammatory form of programmed cell death, has been implicated in eliminating pathogenic infections. However, macrophage pyroptosis-related proteins from <i>Mycobacterium tuberculosis</i> (<i>M.tb</i>) have largely gone unexplored. Here, we identified a cell pyroptosis-inducing protein, Rv1579c, named EST12, secreted from the <i>M.tb</i> H37Rv region of difference 3. EST12 binds to the receptor for activated C kinase 1 (RACK1) in macrophages, and the EST12-RACK1 complex recruits the deubiquitinase UCHL5 to promote the K48-linked deubiquitination of NLRP3, subsequently leading to an NLRP3 inflammasome caspase-1/11-pyroptosis gasdermin D-interleukin-1β immune process. Analysis of the crystal structure of EST12 reveals that the amino acid Y80 acts as a critical binding site for RACK1. An EST12-deficient strain (H37RvΔEST12) displayed higher susceptibility to <i>M.tb</i> infection in vitro and in vivo. These results provide the first proof that RACK1 acts as an endogenous host sensor for pathogens and that EST12-RACK1-induced pyroptosis plays a pivotal role in <i>M.tb</i>-induced immunity.