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. 2021 Sep 17;59(5):86. doi: 10.3892/ijo.2021.5266

Figure 4.

Figure 4

LINC00514 functions as a competing endogenous RNA by sponging miR-378a-5p in ESCC cells. (A) Reverse transcription-quantitative PCR was used to analyze the subcellular localization of LINC00514 in KYSE150 and KYSE30 cells. (B) Subcellular localization of LINC00514 in KYSE150 and KYSE30 cells was investigated using a fluorescence in situ hybridization assay, where LINC00514 was labeled by cyanine 3 (red) and the nuclei were stained with DAPI (blue). (C) lncBase was used to identify the association between LINC00514 and miR-378a-5p. (D) Dual-luciferase reporter assays were performed to investigate the binding between LINC00514 and miR-378a-5p in KYSE150 and KYSE30 cells. An Ago2-RIP assay was used to analyze the enrichment of (E) LINC00514 or (F) miR-378a-5p in the RISC in KYSE150 and KYSE30 cells. (G) Ago2-RIP assay was employed to analyze the enrichment of LINC00514 in KYSE150 and KYSE30 cells in the presence of miR-378a-5p mimic. (H) Transfection of LINC00514 siRNA 2 significantly promoted the expression of miR-378a-5p in KYSE150 and KYSE30 cells. (I) LINC00514 overexpression markedly suppressed the expression of miR-378a-5p in KYSE150 and KYSE30 cells. ****P<0.0001. LINC00514, long intergenic nonprotein-coding RNA 00514; ESCC, esophageal squamous cell carcinoma; miR, microRNA; Ago2, argonaute 2; RIP, RNA immunoprecipitation; RISC, RNA-induced silencing complex; siRNA, small interfering RNA.