circ-PTK2 (hsa_circ_0008305) regulates the pathogenic processes of ovarian cancer via miR-639 and FOXC1 regulatory cascade

• Published in: Cancer cell international 2021-05, Vol.21 (1), p.1-277, Article 277
• Main Authors: Wu, San-Gang, Zhou, Ping, Chen, Jian-Xian, Lei, Jian, Hua, Li, Dong, Yong, Hu, Min, Lian, Chen-Lu, Yang, Li-Chao, Zhou, Juan
• Format: Article
• Language: English
• Subjects: Antibodies; Biomarkers; Cell adhesion & migration; Cell culture; Cell growth; Cell migration; Cell proliferation; Circ-PTK2; DNA helicase; Epithelial–mesenchymal transition; Fluorescence in situ hybridization; FOXC1; Immunofluorescence; Immunohistochemistry; Medical diagnosis; Medical prognosis; Medical screening; Mesenchyme; MicroRNAs; miR-639; miRNA; Ovarian cancer; Polymerase chain reaction; Primary Research; Proteins; Reagents; RNA-directed DNA polymerase; rRNA; siRNA; Western blotting; Wound healing
• ISSN: 1475-2867; 1475-2867
• DOI: 10.1186/s12935-021-01985-x

Background Precise quantification of microRNA is challenging since circulating mRNA and rRNA in the blood are usually degraded. Therefore, it is necessary to identify specific biomarkers for ovarian cancer. This study aimed to investigate candidate circular RNAs (circRNAs) involved in the pathogenic process of ovarian cancer after inhibition of chromodomain helicase DNA binding protein 1-like (CHD1L) and the corresponding mechanism. Methods CHD1L mRNA-targeted siRNA was designed and induced a decreased level of CHD1L function in SK-OV-3 and OVCAR-3 cells observed via transwell and wound healing assays and assessment of epithelial–mesenchymal transition (EMT)-related protein expression by immunofluorescence (IF) and western blotting (WB). After decreasing the level of CHD1L, RNA-seq was conducted, and the circRNA expression profiles were obtained. cirRNAs were then selected and validated by PCR together with Sanger sequencing, fluorescent in situ hybridization (FISH), and reverse transcriptase-quantitative PCR (RT-qPCR). Selected circRNA function in vitro was adjusted via interference and overexpression and assessed via transwell assay, tube formation, and EMT-related protein assay by IF and WB; tumor formation in vivo was followed via hematoxylin and eosin (HE) staining and immunohistochemistry of EMT-related proteins. Based on the competing endogenous RNA prediction of circRNA targets, candidate miRNAs were found, and their downstream mRNAs targeted by the selected miRNA were identified and validated by luciferase assay. The functions of these selected miRNA and mRNA were then further investigated through transwell and WB assay of EMT-related proteins. Results CHD1L was significantly upregulated in ovarian cancer tissues and patients with higher expression of CHD1L had a shorter relapse-free survival (P