The pro-apoptotic factor Bax regulates macrophage necrosis during Mycobacterial infection

Abstract

Abstract In the zebrafish-Mycobacterium marinum model for tuberculosis infection, excess TNF triggers necrosis of infected macrophages which leads to exuberant extracellular bacterial growth and host susceptibility (Tobin et al., 2012; Roca and Ramakrishnan, 2013). It has been shown that in infected macrophages, excess TNF induces production of mitochondrial reactive oxygen species (ROS) which activate two death pathways that converge in macrophage necrosis: cyclophilin D-dependent mitochondrial transition pore and acid sphingomyelinase-dependent ceramide production that leads to lysosomal permeabilization. In exploring the events downstream of lysosomal permeabilization, we found that ceramide acts in a Cathepsin D-dependent manner to induce cell death. In the cytosol Cathepsin D initiates the activation of pro-apoptotic factors including Bid and Bax. We engineered Bax mutants that lack its different functional domains and studied the ability of each mutant protein to regulate apoptosis and necrosis. Then we infected Bax-deficient zebrafish larvae expressing the Bax mutants that were unable to induce apoptosis. We found the BH3 domain, which is required for Bax oligomerization during apoptosis, not to be required for necrosis under excess TNF conditions. Rather, an N-terminal transmembrane helix suggested to be the first portion of Bax in contact with the mitochondrial membrane was found to be required for necrosis. Current studies are focused on understanding post-translational modifications of Bax that determine whether activation of the same protein promotes apoptosis or necrosis.