Alteration in mitochondrial superoxide production by shRNA genesilencing of AIF in various cell lines

Apart from this function in cell death, AIF also plays largely unknown role in cell life. We used AIF gene silencing using shRNA vectors followed by measurements by flow cytometry and confocal fluorescent microscopy to detect the superoxide production during oxidative phosphorylation in mitochondria. We managed to knock-down the expression of gene AIF by transient or stable transfection in various human cell lines. Interestingly, mitochondrial superoxide production in various cell lines reacted to gene silencing of AIF differently. Cell line U-2 OS transiently and also stably transfected with vector shRNA was found to produce less superoxide in comparison to control U-2 OS cells. Interestingly, HeLa cell line shows opposite results with significantly increased mitochondrial superoxide production in silenced HeLa cells. We are also in process of examination of other healthy and cancer cell. Conclusions about possible role of AIF in mitochondria based on our results with various AIF-silenced cell lines will be presented.