FIGURE 2.

Effect of Mdivi-1, Drp1 RNAi, and NAC on blue light-induced mitochondrial fission and Mfn2 expression. (A) Mitochondrial morphology in R28 cells irradiated with or without blue light. Cells were transfected with pDsRed2-Mito to label mitochondria. The fluorescence of mitochondria-targeted DsRed was highly tolerant to prolonged blue light irradiation (please refer to Supplementary Figure 2). The fluorescence signal of pDsRed2-Mito indicated mitochondrial morphology in cells, scale bar = 10 μm. A1′, A2′, A3′, and A4′ show mitochondria with higher magnification in the inserted boxes, scale bar = 5 μm. (B) Effect of Mdivi-1, Drp1 RNAi, and NAC on blue light-induced mitochondrial fission. The average length of mitochondria in each group was measured. The data is presented as the mean ± SD of three independent experiments. 270 mitochondria/group were analyzed. The unit of average length of mitochondria is μm; ^∗∗p < 0.01, ^∗∗∗p < 0.001. (C) Expression level of Mfn2 was immunoblotted in R28 cells after the indicated treatments. β-actin was used as an endogenous control. (D) Relative expression level of Mfn2 was indicated as a normalization of the ratio of Mfn2/β-actin in each sample to the control. The data are presented as the mean ± SD of four independent experiments; ^∗∗p < 0.01; Cont, control; RL, red light; BL, blue light. (E) Silencing efficiency of different siRNAs on Drp1 in R28 cells. Expression level of Drp1 was immunoblotted in R28 cells transfected with siRNA-1∼3 or scramble RNA. Cells without transfection were used as controls. (F) Silencing efficiency of siRNA-3 at 10∼50 nM on Drp1 expression in R28 cells.