Fig. 3
LINC01615 promotes activation of the WNT/β-catenin signaling pathway in GC cells. (A) GSEA of TCGA stomach adenocarcinoma dataset showing enrichment of the WNT/β-catenin signaling pathway in the high-LINC01615 group. (B–C) Western blot analysis of WNT target proteins (TCF4, LEF, Met, c-Myc) in AGS and MKN45 cells after LINC01615 overexpression (B) or knockdown (C), normalized to β-actin. Relative gray value analysis indicates protein levels. (D–E) qRT-PCR analysis of TCF4, LEF, Met, c-Myc mRNA in AGS and MKN45 cells following LINC01615 overexpression (D) or knockdown (E), normalized to GAPDH. (F–H) TOP/FOP luciferase reporter assays showing WNT/β-catenin activity in AGS and MKN45 cells with LINC01615 overexpression (F) or knockdown (G–H). (I–J) Western blot analysis of cytoplasmic and nuclear β-catenin in AGS and MKN45 cells with LINC01615 overexpression (I) or knockdown (J). β-actin and PCNA served as cytoplasmic and nuclear controls, respectively. (K–L) Western blot analysis of β-catenin, p-β-catenin (Ser33/37/Thr41), GSK3β, and p-GSK3β (Ser9) in AGS and MKN45 cells after LINC01615 overexpression (K) or knockdown (L). (M–N) Immunofluorescence staining showing β-catenin localization in AGS (M) and MKN45 (N) cells with LINC01615 overexpression or knockdown. Nuclei were counterstained with DAPI. ns = not significant; *P < 0.05; **P < 0.01; ***P < 0.001; ****P < 0.0001.
