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

Debika Datta,Dhira Rani Saha,Ambika Singh,Gaurav Verma,Shibnath Mazumder,Preeti Khatri,Rajagopal Raman

doi:10.1038/s41420-018-0034-9

Abstract

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 (Ca2+) 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 Ca2+ from ER to mitochondria led to increased mitochondrial Ca2+ (Ca2+)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 (Ca2+)m-uniporter (MUP) blocker ruthenium red, reduced Ca2+ overload suggesting (Ca2+)m fluxes are MUP-driven and the ER-mitochondria tethering orchestrates the process. This is the first report implicating role of sub-cellular Ca2+ in the pathogenesis of M. fortuitum. We summarize, the dynamics of Ca2+ 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.

Highlights

M. fortuitum is a rapidly growing, atypical, nontubercular mycobacteria affecting wide range of animals including humans[1,2,3]
M. fortuitum-induced intracellular Ca2+ imbalance lead to CHOP- mediated headkidney macrophages (HKM) apoptosis Previously, we reported that the imbalance in (Ca2+)c triggers apoptosis in M. fortuitum-infected fish macrophages[24]
The HKM were pre-treated with 2-APB and Dant, specific inhibitors for IP3R and ryanodine receptors (RYRs) respectively[25], infected with M. fortuitum and the changes in CHOP expression and apoptosis studied at 24 h p.i

Summary

Introduction

M. fortuitum is a rapidly growing, atypical, nontubercular mycobacteria affecting wide range of animals including humans[1,2,3]. In fish, it is one of the etiologic agents causing piscine-tuberculosis or mycobacteriosis, a fatal disease characterized by the development of graywhite nodular structures and presence of single or multiple granulomatous lesions on several parenchymal. Official journal of the Cell Death Differentiation Association. Datta et al Cell Death Discovery (2018)4:30 homologous protein (CHOP)[8]. Activation of caspases, a family of cysteine-dependent aspartatedirected proteases, is central to apoptosis and caspase-12 appears to be the prime caspase involved in ER-stress induced apoptosis[18]. Each calpain consists of an 80 kDa catalytic subunit and a common 28 kDa subunit[19]

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Results

Conclusion