ABSTRACT
BACKGROUND & AIMS: Circ-CCT2 (hsa_circ_0000418) is a novel circular RNA that stems from the CCT2 gene. However, the expression of circ-CCT2 and its roles in hepatoblastoma are unknown. Our study aims to study the circ-CCT2 roles in hepatoblastoma development. METHODS: Hepatoblastoma specimens were collected for examining the expression of circ-CCT2, TAF15, and PTBP1. CCK-8 and colony formation assays were applied for cell proliferation analysis. Migratory and invasive capacities were evaluated through wound healing and Transwell assays. The interaction between circ-CCT2, TAF15, and PTBP1 was validated by fluorescence in situ hybridization, RNA pull-down, and RNA immunoprecipitation. SKL2001 was used as an agonist of the Wnt/ß-catenin pathway. A subcutaneous mouse model of hepatoblastoma was established for examining the function of circ-CCT2 in hepatoblastoma in vivo. RESULTS: Circ-CCT2 was significantly up-regulated in hepatoblastoma. Overexpression of circ-CCT2 activated Wnt/ß-catenin signaling and promoted hepatoblastoma progression, whereas knockdown of circ-CCT2 exerted opposite effects. Moreover, both TAF15 and PTBP1 were up-regulated in hepatoblastoma tissues and cells. TAF15 was positively correlated with the expression of circ-CCT2 and PTBP1 in hepatoblastoma. Furthermore, circ-CCT2 recruited and up-regulated TAF15 protein to stabilize PTBP1 mRNA and trigger Wnt/ß-catenin signaling in hepatoblastoma. Overexpression of TAF15 or PTBP1 reversed knockdown of circ-CCT2-mediated suppression of hepatoblastoma progression. SKL2001-mediated activation of Wnt/ß-catenin signaling reversed the anti-tumor effects of silencing of circ-CCT2, TAF15, or PTBP1. CONCLUSIONS: Circ-CCT2 stabilizes PTBP1 mRNA and activates Wnt/ß-catenin signaling through recruiting and up-regulating TAF15 protein, thus promoting hepatoblastoma progression. Our findings deepen the understanding of hepatoblastoma pathogenesis and suggest potential therapeutic targets.
Subject(s)
Hepatoblastoma , Liver Neoplasms , Animals , Mice , Hepatoblastoma/genetics , Hepatoblastoma/pathology , beta Catenin/genetics , beta Catenin/metabolism , RNA, Messenger/genetics , In Situ Hybridization, Fluorescence , RNA/genetics , Liver Neoplasms/genetics , Liver Neoplasms/pathologyABSTRACT
BACKGROUND: Hepatoblastoma is a malignancy that occurs in the liver, most of which occur in children younger than 3 years old. It was reported that lncRNA OIP5-AS1 was up-regulated in hepatoblastoma, but the detailed mechanism by which OIP5-AS1 regulates hepatoblastoma development is unclear. METHODS: qRT-PCR, Western blotting, and immunofluorescence were used to examine levels of OIP5-AS1, PTBP1, ß-catenin or proliferation/stemness-related molecules. Colony formation, sphere formation, wound healing assay and transwell were applied to detect cell proliferation, stemness and invasion, respectively. RIP assay was used to investigate the interaction of OIP5-AS1/PTBP1 and PTBP1/CTNNB1. Finally, in vivo model was constructed to detect the function of OIP5-AS1 in hepatoblastoma. RESULTS: OIP5-AS1 was significantly up-regulated in hepatoblastoma cells. OIP5-AS1 silencing notably attenuated the stemness and invasion of hepatoblastoma cells. OIP5-AS1 bound with PTBP1, and silencing of OIP5-AS1 inhibited ß-catenin. Meanwhile, overexpression of PTBP1 or ß-catenin activation significantly reversed OIP5-AS1 silencing-inhibited hepatoblastoma cell proliferation and stemness. Moreover, ß-catenin was found to be the downstream target of PTBP1, and OIP5-AS1 activated ß-catenin signaling via promoting the binding between PTBP1 and ß-catenin to increase the mRNA stability of ß-catenin. Finally, OIP5-AS1 knockdown significantly alleviated the tumor growth of hepatoblastoma by repressing ß-catenin. CONCLUSION: OIP5-AS1 silencing inhibits the growth and stemness of hepatoblastoma through binding with PTBP1 to inhibit ß-catenin signaling pathway. OIP5-AS1 may be the potential target against hepatoblastoma.
