Fig 2: CCL14-AS interacts with MEP1A mRNA and reduces its stability. A FISH assay was conducted to determine the subcellular location of CCL14-AS in SW620 cells. DAPI‑stained nuclei are blue. Scale bar: 20 μm. B–D RNA fraction assay and online software LncLocator confirmed the subcellular location of CCL14-AS. B Western blot analysis detection of the efficiency of nuclear and cytosolic fractionation. C Nuclear fractionation analyses and qRT-PCR analyses of CCL14-AS expression in the nucleus and cytoplasm. D LncLocator predicts the localization of CCL14-AS in cell. E Gene ontology (GO) analyses of differentially expressed genes from RNA-seq assay of stable CRC cells expressing vector or CCL14-AS. F–G Heatmap (F) and volcanic map (G) showed significant differentially expressed genes after RNA-seq screening. H qRT-PCR confirmed that after overexpressing of CCL14-AS, the mRNA levels of MEP1A decreased, while knockdown of CCL4-AS increased the MEP1A expression. I–J Western blot analysis I and immunofluorescence assay (J) both confirmed that overexpressing CCL14-AS reduced MEP1A protein level. Scale bar: 25 μm. K MS2-RIP experiments detect the direct binding of CCL14-AS to MEP1A mRNA in CRC cells. L RNA-stability analysis showed that CCL14-AS promote the degradation of MEP1A mRNA. ** p < 0.01, *** p < 0.001

Fig 3: CCL14-AS Inhibits CRC invasiveness by down-regulating MEP1A. A, B Western blot (A) and qRT-PCR (B)analyses were performed to examine the expression levels of MEP1A in CCL14-AS overexpressed LoVo and SW620 cells after MEP1A ectopic overexpression. C–E Wound healing assay (C-D) and Migration assays (E) showed that MEP1A ectopic overexpression could prevent the effect of CCL14-AS in inhibiting tumor invasion ability. Scale bar: 100 μm or 50 μm. F, G Popliteal lymph node metastasis model determined MEP1A could reduce the repressive effects of CRC LN metastasis by CCL14-AS. ns: no significance, * p < 0.05, ** p < 0.01, *** p < 0.001

Fig 4: CCL14-AS expression negatively correlates with MEP1A expression in CRC. A qRT-PCR analysis was performed to examine the CCL14-AS and MEP1A mRNA expression in eight human CRC tissues. B Correlation analysis of CCL14-AS and MEP1A at the mRNA (r = 0.857, p = 0.011) in 8 fresh CRC tissues. C The expression of MEP1A in normal intestinal tissues and CRC tissues was examined by IHC. Scale bar: 50 μm. D The expressions of CCL14-AS and MEP1A protein in specimens, including 96 CRC tissue specimens, were examined by FISH and IHC respectively. Scale bar: 100 μm. Representative FISH or IHC images (left) and correlation analysis (right) of CCL14-AS and MEP1A expression. E A working model of CCL14-AS biological function