Fig 2: TGFBR2 was a target of miR-645. (A) Wild-type TGFBR2 3′UTR sequence containing miR-645 binding sites was mutated to generate mutant-type TGFBR2 3′UTR sequence for dual-luciferase reporter assay. (B,C) Dual-luciferase reporter assay was conducted to verify the interaction between miR-645 and TGFBR2. (D) RIP assay was conducted to verify the interaction between miR-645 and TGFBR2. (E,F) The expression of TGFBR2 suppressed by miR-645 restoration was detected by qRT-PCR and western blot. (G,H) The expression of TGFBR2 in tumor tissues (n = 50) and normal tissues (n = 50) was detected by qRT-PCR and western blot. (I,J) The expression of TGFBR2 in LoVo, HCT15 and NCM460 cells was detected by qRT-PCR and western blot. *P < 0.05.

Fig 3: CircFAM120B blocked tumor growth in vivo by regulating miR-645 and TGFBR2. (A) Tumor volume was measured once a week. (B) After 35 days, tumor tissues were removed for weighting. (C) The expression of circFAM120B, miR-645, and TGFBR2 mRNA in removed tumor tissues was detected by qRT-PCR. (D) The expression of TGFBR2 at the protein level was detected by western blot. (E) The enrichment of Ki67 in tumor tissues was detected by immunohistochemical (IHC) staining analysis. *P < 0.05.

Fig 4: miR-645 promoted CRC progression in vitro by mediating TGFBR2. LoVo and HCT15 cells were transfected with anti-miR-645 or anti-miR-645 + si-TGFBR2, with anti-miR-NC or anti-miR-645 + si-NC as the corresponding control. (A,B) The expression of TGFBR2 was examined using qRT-PCR and western blot. (C–E) Cell proliferation was investigated using CCK-8 assay and colony formation assay. (F,G) Cell migration and invasion were monitored by transwell assay (magnification: 100×). (H,I) ECAR was measured by glycolysis stress test. (J,K) Lactate production and glucose consumption were detected using the corresponding kits. (L,M) The expression of HK2 and LDHA was measured by western blot. *P < 0.05.