Study On The Mechanism Of Cell Apoptosis Induced By Down-regulation Mitofilin

Mitofilin is an important mitochondrial protein, which controls the morphylogyof the mitochondrial inner membrane. Down-regulation mitofilin could increase cellapoptosis, but its molecular mechanism is now unclear. We chose mitofilin as a targetto explore the mechanism of down-regulation mitofilin induced cell apoptosis.At first, We explored the function of mitofilin and constructed the lentiviralvector for mitofilin interference. Based on this, we got Hela cells stably inhibitingmitofilin and proceeded further study. We observed mitochondrial net work and itsinner membrane structure by confoncal microscopy and transmission electronmicroscope, respectively. We then used specific dyes labling different cellularmolecules to test mitochondrial function by FACS anlysis. We found that inhibitingmitofilin expression could lead to a series of mitochondrial disfunction.Under normal condition, down-regulation mitofilin could increase cell apoptosisin normal condition. So our study detected cell apoptosis after over and down-expressmitofilin after etoposide (10μg/mL) stimuli. Our previous work found the interationbetween Tim23and mitofilin by two yeast-hybrid and GST-pull down technology.This work proved the interation between mitofilin and Tim23by both exogenous andendogenous Co-IP experiment. Further truncated mitofilin and Tim23constructionand GST-pull down study showed that the exactly interating domain between the twoproteins. We further explored the effects of cell apoptosis after overexpression Tim23and its truncated protein lack of mitofilin interacting domain.Our data also showed that overexpression mitofilin could effectively inhibitTim23degradation and reduce cell apoptosis after stimuli of etoposide. In contrast,down-regulation mitofilin could promote Tim23degradation and accelerate apoptosisprocess. Overexpress Tim23could significantly reduce down-regulation mitofilininduced cell death, but the truncated Tim23couldn’t.We concluded that mitofilin is involved cell apoptosis via interacting with Tim23.Our research founded the mechanism of mitofilin on cell apoptosis and established thefundament for further exploring the biological effects mediated by this interaction.