Mitochondrial inner membrane protein (mitofilin) knockdown induces cell death by apoptosis via an AIF-PARP-dependent mechanism and cell cycle arrest

Mitofilin is an inner membrane protein that has been defined as a mitochondria-shaping protein in controlling and maintaining mitochondrial cristae structure and remodeling. We determined the role of mitofilin in cell survival by investigating the mechanism underlying mitofilin knockdown-induced cell death by apoptosis. Cultured H9c2 myoblasts and HEK 293 cells were treated with mitofilin siRNA or scrambled siRNA for 24 hours. Cell death (apoptosis), caspase 3 activity, and cell cycle phases were assessed by flow cytometry, while cytochrome C release and intracellular ATP production were measured by ELISA. Mitofilin, AIF and PARP expression were measured by Western blot analysis and calpain activity assessed using a calpain activity kit. Mitochondrial images were taken using electron microscopy. We found that mitofilin knockdown increases apoptosis mainly via activation of the AIF-PARP pathway leading to nuclear fragmentation that is correlated with S phase arrest of the cell cycle. Knockdown of mitofilin...