Fig 1: Circ_0058124 knockdown blocked tumor development in vivo. TPC-1 cells transfected with sh-circ_0058124 or sh-NC were inoculated into nude mice. (A) Tumor volume was recorded once a week after inoculation. (B) Tumor weight was measured in each group after 28 days when tumor tissues were excised. (C and D) The expression of circ_0058124 and miR-370-3p in the removed tissues was detected by qRT-PCR. (E) The expression of LMO4 at the protein level was detected by Western blot. *P<0.05.
Fig 2: In situ proximity ligation assay with the SH-SY5Y and Mut5 cells treated, or not, with leptomycin B (L). In (a), representative pictures of BRCA1-BARD1 heterodimers (red dots) showing higher numbers in the cytoplasm and the nuclei (DAPI staining) of the Mut5 cells compared to the wild-type cells. Scale bar: 75 µM. In (b), graphical representation of the statistical analyses of the experiment for BRCA1-BARD1 heterodimers (n = 9). The data analyses for the other heterodimers, BRCA1-LMO4, Fe65-Tip60, and Fe65-pTip60, are also shown (n = 9). We observed statistically significant differences (ANOVA with Sidak's multiple comparison test: *p < 0.05; **p = 0.002; #p = 0.0002) between both strains in the cytoplasm (CYTO), not in the cell nuclei. The nuclei are labelled NU+CHR according to the denomination used for the Western blots (NU: nucleoplasm; CHR: chromatin).
Fig 3: Linc-ROR promotes HNSCC cell proliferation and invasion via LMO4-mediated AKT/PI3K pathway activation. (A) The detection of Linc-ROR and LMO4 expression by RT-qPCR in TSCCA cells upon overexpression or silencing of LMO4 or Linc-ROR, sh-Linc-ROR + oe-LMO4, LY294002 or oe-LMO4 + LY294002. *p < 0.05, compared with the oe-NC group, # p < 0.05, compared with the sh-NC group; & p < 0.05, compared with the oe-LMO4 group; (B) Protein expression of LMO4, AKT, and PI3K along with p-AKT and p-PI3K in TSC7CA cells upon overexpression or silencing of LMO4 or Linc-ROR, sh-Linc-ROR + oe-LMO4, LY294002 or oe-LMO4 + LY294002. *p < 0.05, compared with the oe-NC group, # p < 0.05, compared with the sh-NC group; & p < 0.05, compared with the oe-LMO4 group; @ p < 0.05, compared with the LY294002 group; (C) Proliferation of TSCCA cells measured by monoclonal formation assay upon overexpression or silencing of LMO4 or Linc-ROR, sh-Linc-ROR + oe-LMO4, LY294002 or oe-LMO4 + LY294002; *p < 0.05, compared with the oe-NC group, # p < 0.05, compared with the sh-NC group; & p < 0.05, compared with the oe-LMO4 group; @ p < 0.05, compared with the LY294002 group; (D) Proliferation of TSCCA cells measured by MTT assay upon overexpression or silencing of LMO4 or Linc-ROR, sh-Linc-ROR + oe-LMO4, LY294002 or oe-LMO4 + LY294002. *p < 0.05, compared with the oe-NC group, # p < 0.05, compared with the sh-NC group; & p < 0.05, compared with the oe-LMO4 group; @ p < 0.05, compared with the LY294002 group; (E), Invasion of TSCCA cells measured by Transwell assay upon overexpression or silencing of LMO4 or Linc-ROR, sh-Linc-ROR + oe-LMO4, LY294002 or oe-LMO4 + LY294002. *p < 0.05, compared with the oe-NC group, # p < 0.05, compared with the sh-NC group. & p < 0.05, compared with the oe-LMO4 group; @ p < 0.05, compared with the LY294002 group. All experiments were repeated three times.
Fig 4: Silencing of Linc-ROR reverses the promoting effect of overexpressing FOXM1 on tumor growth in vivo. (A) Tumor growth curve; (B) Tumor size and weight; *p < 0.05, compared with the oe-NC group, # p < 0.05, compared with the sh-NC group; (C) Expression of FOXM1, Linc-ROR, and LMO4 in mouse tumor tissues determined by RT-qPCR; *p < 0.05, compared with the oe-NC group, # p < 0.05, compared with the sh-NC group; (D) Protein expression of FOXM1 and LMO4 in mouse tumor tissues detected by Western blot; *p < 0.05, compared with the oe-NC group, # p < 0.05, compared with the sh-NC group; & p < 0.05, compared with the oe-FOXM1 group; (E) The positive expression of FOXM1, LMO4, and Ki-67 proteins in mouse tumor tissues detected by IHC; (F) The apoptosis of cells in mouse tumor tissues detected by TUNEL assay. n = 15 for mice per group.
Fig 5: MiR-370-3p bound to LMO4 3'UTR and suppressed LMO4 expression. (A) LMO4 was a putative target of miR-370-3p, which was predicted by the online tool starbase. (B and C) The association between miR-370-3p and LMO4 was verified by dual-luciferase reporter assay. (D) The expression of LMO4 in PTC tissues (n=20) and normal tissues (n=20) was detected by qRT-PCR. (E) The expression of LMO4 in Nthy-ori 3–1, IHH-4 and TPC-1 cells was measured by Western blot. (F) The efficiency of LMO4 overexpression was detected by Western blot. (G) The expression of LMO4 in IHH-4 and TPC-1 cells transfected with miR-370-3p mimic, miRNA NC, miR-370-3p mimic+pc-LMO4 or miR-370-3p mimic+pc-NC was detected by Western blot. (H) The expression of LMO4 in IHH-4 and TPC-1 cells transfected with si-circ_0058124, si-NC, si-circ_0058124+miR-370-3p inhibitor or si-circ_0058124+inhibitor NC was quantified by Western blot. *P<0.05.
Supplier Page from Abcam for Anti-LMO4 antibody