CircFOXK2 Promotes Tumor Growth and Metastasis of Pancreatic Ductal Adenocarcinoma via Complexing with RNA Binding Proteins and Sponging MiR-942

Abstract

Abstract Objective Circular RNA (circRNA) is a novel class of non-coding RNAs that regulate gene expression. However, the role of circRNAs in pancreatic ductal adenocarcinoma (PDAC) is largely unknown. Design We performed circRNA sequencing of non-tumor HPDE and PDAC cells. We investigated the functions of circFOXK2 in PDAC by gain-of-function and loss-of-function assays. Bioinformatics analysis, luciferase assay and microRNA pulldown assays were performed to identify circFOXK2 interacting-miRNAs. To further investigate the mechanism, we performed circRNA-pulldown and mass spectrometry to identify circFOXK2-interacting proteins in PDAC. Results We identified 169 differentially expressed circRNAs in PDAC cells. We validated that one of the circRNAs circFOXK2 was significantly up-regulated in PDAC cells and in 63 % of primary tumor (53 out of 84). Gain-of-function and loss-of-function assays demonstrated that circFOXK2 promoted PDAC cell growth, migration and invasion. CircFOXK2 was also involved in cell cycle progression and apoptosis. circFOXK2 functioned as sponge for miR-942, and in turn promoted the expression of miR-942 targets ANK1, GDNF and PAX6. Furthermore, circFOXK2 interacted with 94 proteins, which were involved in cell adhesion and mRNA splicing. Among these circFOXK2-interacting proteins, YBX1 and hnRNPK were validated by RNA immunoprecipitation. Importantly, circFOKX2 interacted with YBX1 and hnRNPK targets NUF2 and PDXK in PDAC cells. Knockdown of circFOXK2 reduced the binding of YBX1 and hnRNPK to NUF2 and PDXK, and in turn decreased their expressions in PDAC cells. Conclusion We identified that circFOXK2 promoted PDAC cells growth and metastasis. Also, circFOXK2 complexed with YBX1 and hnRNPK to promote the expressions of oncogenic proteins. Significance of this study What is already known on this subject? Differentially expressed circRNAs are involved in carcinogenesis of many cancers. CircRNAs function as microRNA sponges to regulate gene expression. The roles of circRNAs in PDAC progression is largely unknown. What are the new findings? circFOXK2 is upregulated in PDAC primary tumors. circFOXK2 promotes PDAC tumor growth and liver metastasis. circFOXK2 functions as sponges for miR-942 to promote the expressions of oncogenic ANK1, GDNF and PAX6. circFOXK2 complexes with YBX1 and hnRNPK to promote the expressions of oncogenic proteins in PDAC. How might it impact on clinical practice in the foreseeable future? circFOXK2 upregulation in PDAC may function as a novel biomarker for diagnosis. circFOXK2 may be a novel therapeutic target in treating PDAC.