Figure 5.

Circular ANRIL species participate in INK4 gene regulation in proliferative or senescent cells

(A) Senescent WI38 hTERT RAF1-ER cells were transfected with the indicated siRNA. Seventy-two hours later, total RNA was extracted. ANRIL, p15, p16, p14^ARF and p21 mRNA expression was measured by RT-qPCR using the primers indicated in parentheses, calculated relative to GAPDH and normalized to 1 for the control siRNA in each experiment. The means and standard deviations from 3 to 14 independent experiments are shown, depending on the primers. Significant differences are indicated with asterisks (*: p value < 0.05, ** to ****: p values < 10⁻², 10⁻³ and 10⁻⁴, respectively, two-sided paired Student’s t-test on log2 values); the number of the p value is indicated when it is between 0.05 and 0.1; ns: not significant. (B) Same as in (A), except that the indicated siRNAs were used. (C) Proliferative WI38 hTERT RAF1-ER cells were transfected using the indicated siRNAs and analysed as in (A). (D) Senescent WI38 hTERT RAF1-ER cells were transfected with the indicated siRNAs. 24 to 72 h later, the cells were transfected with the indicated in vitro synthesized circular RNAs. 24 h later, total RNA was extracted and analysed by RT-qPCR using the primers indicated in parentheses. The means and standard deviations from four independent experiments are shown. Significant differences are indicated as in (A).