Mycobacterium fortuitum-induced ER-Mitochondrial calcium dynamics promotes calpain/caspase-12/caspase-9 mediated apoptosis in fish macrophages

Mycobacterium fortuitum is a natural fish pathogen. It induces apoptosis in headkidney macrophages (HKM) of catfish, Clarias sp though the mechanism remains largely unknown. We observed M. fortuitum triggers calcium (Ca(2+)) insult in the sub-cellular compartments which elicits pro-apototic ER-stress factor CHOP. Alleviating ER-stress inhibited CHOP and attenuated HKM apoptosis implicating ER-stress in the pathogenesis of M. fortuitum. ER-stress promoted calpain activation and silencing the protease inhibited caspase-12 activation. The study documents the primal role of calpain/caspase-12 axis on caspase-9 activation in M. fortuitum-pathogenesis. Mobilization of Ca(2+) from ER to mitochondria led to increased mitochondrial Ca(2+) (Ca(2+))(m) load,(,) mitochondrial permeability transition (MPT) pore opening, altered mitochondrial membrane potential (ΔΨm) and cytochrome c release eventually activating the caspase-9/-3 cascade. Ultra-structural studies revealed close apposition of ER and mitochondria and pre-treatment with (Ca(2+))(m)-uniporter (MUP) blocker ruthenium red, reduced Ca(2+) overload suggesting (Ca(2+))(m) fluxes are MUP-driven and the ER-mitochondria tethering orchestrates the process. This is the first report implicating role of sub-cellular Ca(2+) in the pathogenesis of M. fortuitum. We summarize, the dynamics of Ca(2+) in sub-cellular compartments incites ER-stress and mitochondrial dysfunction, leading to activation of pro-apoptotic calpain/caspase-12/caspase-9 axis in M. fortuitum-infected HKM.