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. 2021 May 24;11(15):7322–7336. doi: 10.7150/thno.56990

Figure 5.

Figure 5

ECRRs promote the endogenous circular RNAs production. (A-B) The schematic diagrams of how ECRRs modulate the production of the endogenous circular RNA circ10720 (A) or circBIRC6 (B). (C) ECRR (PUF10720-L-GTPase) and ECRR (PUF10720-R-GTPase) were stably transfected into 293T cells. The resulting cells were transiently transfected with circ10720 siRNA or control siRNA. After forty-eight hours, total RNAs were isolated from the transfected cells, and qRT-PCR assay was applied to examine the levels of circ10720 and linear RNA (CUL2 E4-E5). Three experiments were carried out with mean +/- SD of relative fold change of circ10720 and linear Cul2 expression plotted. (D) ECRR (PUF10720-L-BTB) and ECRR(PUF10720-R-BTB) were stably transfected into 293T cells. The resulting cells were transiently transfected with circ10720 siRNA or control siRNA. After forty-eight hours, total RNAs were isolated from the transfected cells, and qRT-PCR assay was applied to examine the levels of circ10720 and linear RNA (CUL2 E4-E5). Three experiments were carried out with mean +/- SD of relative fold change of circ10720 and linear Cul2 expression plotted. (E) ECRR (PUFBIRC6-L-GTPase) and ECRR (PUFBIRC6-R-GTPase) were stably transfected into 293T cells. The resulting cells were transiently transfected with circBIRC6 siRNA or control siRNA. After forty-eight hours, total RNAs were isolated from the transfected cells, and qRT-PCR assay was applied to examine the levels of circBIRC6 and linear BIRC6 RNA. Three experiments were carried out with mean +/- SD of relative fold change of circBIRC6 and linear BIRC6 expression plotted. (F) ECRR (PUFBIRC6-L-BTB) and ECRR (PUFBIRC6-R-BTB) were stably transfected into 293T cells. The resulting cells were transiently transfected with circBIRC6 siRNA or control siRNA. After forty-eight hours, total RNAs were isolated from the transfected cells, and qRT-PCR assay was applied to examine the levels of circBIRC6 and linear BIRC6 RNA. Three experiments were carried out with mean +/- SD of relative fold change of circBIRC6 and linear BIRC6 expression plotted. For panels C to J, * indicates p < 0.05, ** indicates p < 0.01, *** indicates p < 0.001. (G-H) ECRRs (PUF10720-L, R-GTPase) or ECRRs (PUF10720-L, R-BTB) were stably transfected into HeLa (G) or H1299 (H) cells. Total RNAs were isolated from the transfected cells, and qRT-PCR assay was applied to examine the levels of circ10720 and linear RNA (CUL2 E4-E5). Three experiments were carried out with mean +/- SD of relative fold change of circ10720 and linear Cul2 expression plotted. (I-J) ECRRs (PUFBIRC6-L, R-GTPase) or ECRRs (PUFBIRC6-L, R-BTB) were stably transfected into HeLa (I) or H1299 (J) cells. Total RNAs were isolated from the transfected cells, and qRT-PCR assay was applied to examine the levels of circBIRC6 and linear BIRC6 RNA. Three experiments were carried out with mean +/- SD of relative fold change of circBIRC6 and linear BIRC6 expression plotted. For panels C to J, * indicates p < 0.05, ** indicates p < 0.01, *** indicates p < 0.001.