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. 2020 May 21;15(5):e0225356. doi: 10.1371/journal.pone.0225356

Fig 1. MiR-148a regulates (P)RR expression by targeting its 3’-UTR sequences.

Fig 1

A. An illustration showing that there are two predicted binding sites of miR-148a on the 3’-UTR of human (P)RR. HepG2 and Huh7 cells, cultured with serum containing medium, were transfected with miR-148a or control miRNA for 48 hours, and gene expression and protein abundances were analyzed. B. (P)RR mRNA level was determined by quantitative PCR, and corrected for 36B4 level in the same sample and expressed as ratio of the control miRNA transfected. Results are from four independent experiments in triplicates. N = 12; ***: p<0.001. C. Total cell lysates were blotted as indicated and a representative blot of 3 independent experiments in triplicates was shown. D. (P)RR protein abundance was quantified and normalized to the level of tubulin in the same lysates, and expressed as the relative ratio of (P)RR abundance in control miRNA transfected. N = 9; ***: P<0.001. E. An illustration showing two constructs (Mut1 and Mut2) which are mutated for the binding site for miR-148a on the 3’-UTR of human (P)RR, comparing to wildtype (WT) sequence. F. HEK293T cells were transfected with luciferase reporter plasmids constructed using wildtype (WT) and mutated (Mut1 and Mut2) 3’-UTR of human (P)RR, together with either control miRNA or miR-148a. Cells were cultured with serum containing medium. Firefly luciferase activity was measured and corrected for Renilla luciferase activity in the same sample, and expressed as ratio of WT reporter plasmid transfected samples. Results are from four independent experiments in triplicates (N = 12). ###: WT+miR-148a vs. WT+control miRNA, p<0.001; N.S (not significant): Mut1+miR-148a vs. WT+miR-148a; &&&: Mut2+miR-148a vs. WT, p<0.001.