Mtln expression was inhibited in cultured human HeLa cells using siRNAs, and protein measures and mitochondrial imaging analyses were performed 48 hr later.
(A) Western blot shows clear reductions in Mtln expression in cells treated with Mtln-targeted siRNAs (siMtln1–4, each representing unique siRNA sequences), relative to two non-targeted negative control siRNAs (siNC1–2). 10 nM (top) and 50 nM (bottom) doses were tested. OXPHOS cocktail protein levels did not change in response to Mtln knockdown in any siMtln treatment group.
(B) Top, representative photomicrographs depicting TMRE (mitochondrial membrane potential) and MitoSOX (ROS) probe intensities in HeLa cells treated with 50 nM of the indicated siRNAs. Scale bar, 20 μm. Bottom, quantified probe intensities are plotted as mean ± SEM, sample n is indicated within each bar, and p values were determined by one-way ANOVA with Dunnett’s post hoc (****p < 0.0001 compared to βgal+Dox).