Fig 2: Sec62 induces UCA1 expression via MAPK signalling pathway. (A) In Sec62 overexpressing and knockdown cell models, total and phosphorylation of P38, ERK, JNK and p‐ATF2 protein expression were detected by western blot. (B) SW480 cell was transfected with Sec62 shRNA or shNC lentivirus and then treated with JNK agonist Anisomycin or ATF2 siRNA, the metastatic capability of cells was determined by trans‐well (up) and wound‐healing assays (below). The percentage of wound gap was normalized to 0 h. Scale bars, 10 μm. (C) qRT‐PCR outcomes of UCA1 levels in the indicated cells. (D) Trans‐well (up) and wound healing (below) assays measured the metastatic ability of HCT116 cells transfected with LV‐Sec62 or LV‐NC lentivirus and then treated with JNK inhibitor SP600125. The percentage of wound gap was normalized to 0 h. Scale bars, 10 μm. (E) qRT‐PCR analysis of UCA1 expression in the indicated cells. **p < 0.01 and *p < 0.05, N ≥ 3. Data are presented as mean ± SD

Fig 3: Sec62 is upregulated in colorectal cancer (CRC) and predicts poor overall survival. (A and B) qRT‐PCR and western blot, respectively shownthe mRNA and protein level of Sec62 in different CRC cell lines (n = 12) and FHC, an immortalized colonic epithelial cell line. (C and D) The expression of Sec62 in paired primary CRC tissues (T, n = 30) and adjacent non‐tumour samples (N, n = 30) was determined by Western blot and qRT‐PCR analysis. (E) The transcriptional level of Sec62in the CRC primary site and metastatic site was identified from GEO database (GSE41258, GSE68468, GSE18105). (F) CPTAC database was used to detect the protein level of Sec62 between CRC tissues and normal colon tissues. (G) Representative positive immunohistochemical (IHC) staining for Sec62 of paired CRC (n = 100) and adjacent non‐tumour samples (n = 80) (left). The semiquantitative analysis evaluated the scores of Sec62 expression in samples (right). Scale bars, 100 μm (up) or 20 μm (below). (H) Kaplan–Meier analysis exhibited the different overall survival (OS) time in patients with CRC with different Sec62 level. ***p < 0.001, **p < 0.01 and *p < 0.05. Each assay was repeated thrice (N ≥ 3), Data are presented as mean ± standard deviation (SD)

Fig 4: UCA1 is indispensable for Sec62‐mediated CRC cell metastasis. (A) Volcano map exhibited differentially expressed genes when Sec62 was inhibited in DLD‐1 cells (Sig‐UP: significant up‐regulation; No‐Diff: no difference; Sig‐Down: significant down‐regulation). (B) qRT‐PCR analysis of obvious changed candidate genes in Sec62‐silencing DLD‐1 cells. (C) The expression of UCA1 was detected by qRT‐PCR in Sec62 upregulated or downregulated CRC cells. (D) The level of UCA1 in paired CRC tissues and non‐tumour tissues was determined by qRT‐PCR (Left, n = 30). A positive correlation between the mRNA level of Sec62 and UCA1 in CRC tissues was identified (right, n = 30). (E) GEO datasets was utilized to analyse UCA1 transcriptional level among healthy donor colon tissues (healthy donor, n = 50), paired non‐tumour normal colon tissues (paired normal, n = 98) and CRC tissues (tumour tissue, n = 98) (left, GSE44076); and the level of UCA1 in primary sites (n = 39) and metastatic sites (n = 94) was measured from GSE41568 (right). (F) UCA1 expression in CRC cell lines (n = 10) and two immortalized colon epithelial cell lines, FHC and NCM460 was identified by qRT‐PCR. (G and H) Trans‐well (G) and wound healing (H) assays shown the motility ability of Sec62 silencing DLD‐1 cells after transfection with plasmid of UCA1 (left, Control = control vector, UCA1 = UCA1 pcDNA3.1) and Sec62 overexpressing HCT116 cells transfected with of siRNA UCA1 (right). The percentage of wound gap was normalized to 0 h. Scale bars, 10 μm. **p < 0.01 and *p < 0.05, ns = No Significance. N ≥ 3. Data are presented as mean ± SD

Fig 5: Schematic illustration of the Sec62/MAPK/ATF2 /UCA1 axis in metastatic colorectal cancer cells