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. 2021 Aug 2;11:699933. doi: 10.3389/fonc.2021.699933

Figure 6.

Figure 6

EIF4A3 downregulates the expression of CASC2 via inducing its cleavage, while RORA transcriptionally regulates the expression of EIF4A3. (A–D) qPCR (A, C) and western blotting (B, D) showing the validation of EIF4A3 knockdown or overexpression. (E, F) The RNA immunoprecipitation (RIP) assay was performed in GSC4D or GSC2C after EIF4A3 knockdown or overexpression, and negative control was transfected, followed by q-PCR to detect the enrichment of CASC2 and EIF4A3. (G, H) The RNA pull-down assays showed the EIF4A3 protein immunoprecipitation with CASC2 as detected by western blotting. (I) RT-qPCR showing the expression of CASC2 after EIF4A3 knockdown or overexpression. (J) The nascent CASC2 was detected by RT-qPCR. (K) Graphs showing CASC2 levels at different times treated by actinomycin D in GSC4D and GSC2C. (L) RT-qPCR showing the expression of EIF4A3 after RORA knockdown or overexpression. (M) Western blotting showing the expression of EIF4A3 protein in RORA-silenced GSC2C and RORA-overexpressed GSC4D. (N) The luciferase reporter assays showed that RORA knockdown or overexpression affected the luciferase activities of EIF4A3 in GSCs. EV, empty vector; OE, overexpression; NC, negative control; KD, knockdown. All data are expressed as the mean ± SD (three independent experiments). **P < 0.01; ***P < 0.001; n.s., not significant.