Fig. 4

HnRNPC promotes XB130 expression post-transcriptionally by binding to XB130 mRNA 3’UTR. A-B: RT-qPCR and Western blotting were conducted to assess the expression of XB130 in control and hnRNPC-silenced (A) or -overexpressing (B) cells. GAPDH was used as an internal control. C: A dual-luciferase reporter recombinant vector (PEZX-XB130 promoter) containing the XB130 promoter was constructed and transfected into control and hnRNPC-silenced cells. Dual-luciferase activity was measured 48 h post-transfection. hRluc was used as an internal control. D: The wild-type full-length XB130 3’UTR (3’UTR), along with the 3’UTR segments 113–230 and 503–660, and a mutated XB130 3’UTR lacking the segments 113–230 and 503–660 (MU-3’UTR), were generated through in vitro transcription. RNA pull-down assays were performed to investigate the interactions of these RNA fragments with hnRNPC and Hsp90 in NCI-H292 cells. E: RIP assays verified the interactions of hnRNPC protein with XB130 and Hsp90 mRNA in hnRNPC-overexpressing cells. IgG-bound RNA served as a negative control. F: A series of dual-luciferase reporter recombinant vectors containing different segments of XB130 mRNA 3’UTR were constructed and transfected into control and hnRNPC-silenced cells. Dual-luciferase activity was assessed 48 h after transfection. hluc served as an internal control. * p < 0.05, ** p < 0.01, *** p < 0.001