RT Journal Article SR Electronic(1) A1 Lu, Jia-Rong A1 Lu, Wen-Wen A1 Lai, Jian-Zhong A1 Tsai, Fu-Lian A1 Wu, Szu-Hsien A1 Lin, Cheng-Wen A1 Kung, Szu-HaoYR 2013 T1 Calcium flux and calpain-mediated activation of the apoptosis-inducing factor contribute to enterovirus 71-induced apoptosis JF Journal of General Virology, VO 94 IS 7 SP 1477 OP 1485 DO https://doi.org/10.1099/vir.0.047753-0 PB Microbiology Society, SN 1465-2099, AB Enterovirus 71 (EV71) is a causative agent of an array of childhood diseases with severe neurological manifestations implicated. EV71 infection is known to induce caspase-dependent apoptosis in cell cultures and animal models. However, whether an alternative apoptotic pathway independent of caspase activation can be triggered by EV71 infection has not been explored. In this study, we showed that calcium (Ca2+)-activated calpains are capable of mediating caspase-independent pathway activation during EV71-induced apoptosis in HeLa cells. Results from subcellular fractionation analysis and confocal imaging indicated that during EV71 infection, apoptosis-inducing factor (AIF), a primary mediator of the caspase-independent pathway, became truncated and translocated from the mitochondrion to nucleus. This was accompanied by the release of cytochrome c, and sharply decreased mitochondrial membrane potential. AIF knockdown data indicated significant protection against apoptotic cell death, with greater protection provided by the addition of a pan-caspase inhibitor. The Ca2+-dependent, calpain isoforms 1 and 2, but not cathepsins, were proven crucial for the altered AIF behaviour as studied by the pharmacological inhibitor and the knockdown approaches. We then analysed Ca2+ dynamics in the infected cells and found elevated levels of mitochondrial Ca2+. Treatment with ruthenium red, a mitochondrial Ca2+ influx inhibitor, significantly blocked calpain activations and AIF cleavage. Our conclusion was that calpain activation via Ca2+ flux plays an essential role in eliciting an AIF-mediated, caspase-independent apoptotic pathway in EV71-infected cells. These findings should be useful for understanding the virus-induced cytopathology and the impact of Ca2+ homeostasis on EV71 infection., UL https://www.microbiologyresearch.org/content/journal/jgv/10.1099/vir.0.047753-0