Increasing evidence has implicated that circular RNAs (circRNAs) exert important roles in colorectal cancer (CRC) occurrence and progression. However, the role of a novel circRNA, circUHRF2, remains unknown in CRC. Our work aimed at identifying the functional roles of circUHRF2 in CRC and illustrating the potential mechanisms. As assessed by quantitative real-time PCR (qRT-PCR), circUHRF2 and methyltransferase-like 3 (METTL3) were highly expressed in CRC specimens and cells. Sanger sequencing and RNase R assays were performed to verify the ring structure of circUHRF2. Notably, aberrantly increased expression of circUHRF2 was positively correlated with poor prognosis of CRC patients. Functional experiments indicated that CRC stemness, migration, and epithelial-mesenchymal transition (EMT) were suppressed by the knockdown of circUHRF2 or METTL3. Mechanistically, METTL3 enhanced circUHRF2 expression through N6-methyladenine (m⁶A) modification. Rescue experiments showed that overexpression of circUHRF2 reversed the repressive effect of METTL3 silencing on CRC progression. Moreover, circUHRF2 inhibited the loss of DEAD-box helicase 27 (DDX27) protein via promoting the interaction between insulin-like growth factor 2 mRNA-binding protein 1 (IGF2BP1) and DDX27 mRNA. DDX27 knockdown repressed CRC malignant properties, which was counteracted by circUHRF2 overexpression. The in vivo assays in nude mice demonstrated that circUHRF2 or METTL3 silencing exerted a suppressive effect on CRC growth and liver metastasis via repressing DDX27 protein expression. Taken together, METTL3-mediated m⁶A modification upregulated circUHRF2 and subsequently inhibited loss of DDX27 protein via recruitment of IGF2BP1, which conferred CRC stemness and metastasis. These findings shed light on CRC pathogenesis and suggest circUHRF2 as a novel target for CRC treatment.

Background: Colorectal cancer (CRC) is the third frequently diagnosed cancer with high incidence and mortality rate worldwide. Our previous report has demonstrated that circCOL1A1 (hsa_circ_0044556) functions as an oncogene in CRC, and Gene Ontology (GO) analysis has also revealed the strong association between circCOL1A1 and angiogenesis. However, the mechanism of circCOL1A1 or exosomal circCOL1A1 in CRC angiogenesis remains elusive. Methods: Purified exosomes from CRC cells were characterized by nanoparticle tracking analyzing, electron microscopy and western blot. qRT-PCR, immunohistochemistry or western blot were employed to test the expression of circCOL1A1, EIF4A3, Smad pathway and angiogenic markers. Cell proliferation of HUVECs was monitored by CCK-8 assay. The migratory and angiogenic capabilities of HUVECs were detected by wound healing and tube formation assay, respectively. Bioinformatics analysis, RNA immunoprecipitation (RIP), RNA pull-down and FISH assays were used to detect the interactions among circCOL1A1, EIF4A3 and Smad2/3 mRNA. The in vitro findings were verified in xenograft model. Results: CRC cell-derived exosomal circCOL1A1 promoted angiogenesis of HUVECs via recruiting EIF4A3. EIF4A3 was elevated in CRC tissues, and it stimulated angiogenesis of HUVECs through directly binding and stabilizing Smad2/3 mRNA. Moreover, exosomal circCOL1A1 promoted angiogenesis via inducing Smad2/3 signaling pathway in vitro, and it also accelerated tumor growth and angiogenesis in vivo. Conclusion: CRC cell-derived exosomal circCOL1A1 promoted angiogenesis via recruiting EIF4A3 and activating Smad2/3 signaling.

Background: Colorectal cancer (CRC) is one of the most commonly diagnosed cancers with high incidence and poor prognosis worldwide. Circ_RNF13 is upregulated in CRC; however, the biological roles and downstream signaling of circ_RNF13 remain undefined. Methods: The characterization of circ_RNF13 was determined by Sanger sequencing, qRT-PCR, subcellular fractionation assay, and RNA FISH. Western blot analysis and qRT-PCR were employed to detect the expression of the key molecules and stemness markers in CRC tumor samples and cells. The stem-like activities of CRC cells were assessed by sphere formation assay, flow cytometry, and immunofluorescence (IF). Cell viability was monitored by CCK-8 assay. The chemosensitivity of CRC cells was assessed by colony formation and cell apoptosis assays. Bioinformatics analysis, RIP assay, RNA pull-down assay, and FISH/IF staining were used to detect the association between circ_RNF13 and TRIM24. The transcriptional regulation of DDX27 was investigated by ChIP assay, and the post-translational regulation of TRIM24 was detected by Co-IP. The in vitro findings were verified in a xenograft model. Results: circ_RNF13 and DDX27 were elevated in CRC tumor samples and cells. Knockdown of circ_RNF13 or DDX27 inhibited stemness and increased chemosensitivity in CRC cells. Mechanistically, circ_RNF13 regulated DDX27 expression via TRIM24-mediated transcriptional regulation, and circ_RNF13 stabilized TRIM24 via suppressing FBXW7-mediated TRIM24 degradation. In vivo studies revealed that the knockdown of circ_RNF13 impaired stemness and enhanced the chemosensitivity of CRC in the xenograft model. Conclusion: circ_RNF13 regulated the stemness and chemosensitivity of CRC by transcriptional regulation of DDX27 mediated by TRIM24 stabilization.