Fig. 1

Regulation of PINK1 by miRNA. a Schematic diagram of AGO-specific RNA immunoprecipitation assay. b, c PINK1 mRNAs are associated with miRNA/RISC complex. HeLa cells were transfected with 50 nM of AGO2 siRNA or negative control. 48 h post-transfection, AGO2-bound mRNAs were pulled down from cell lysates with 2A8 anti-AGO2 antibody or non-specific IgG antibody. PINK1 mRNA levels were then measured by qRT-PCR. siRNA against AGO2 served as a quality control for AGO-specific RNA immunoprecipitation assay. PINK1 mRNA levels were quantified as a percentage of group 1 (n = 6, two-way ANOVA). d, e Knock-down of DICER1 increased PINK1 protein levels. HeLa cells were transfected with 50 nM DICER1 siRNA (DICER1 KD) or negative control (Ctl). 48 h post-transfection, cells were harvested for Western blot. The arrowhead indicates a full-length PINK1 band and the asterisk indicates a non-specific band. Each protein level was normalized to corresponding GAPDH level and quantified as a percentage of control (n = 4, t-test). Values are mean ± SEM (n.s. = non-significant, **p < 0.01, ***p < 0.001)