Expression profiling of circular RNA reveals a potential miR-145-5p sponge function of circ-AFF2 and circ-ASAP1 in renal cell carcinoma.

objectivesCircular RNAs (circRNAs) are involved in carcinogenesis, though their expression profile in renal cell carcinoma (RCC) is uncharacterized. The tumor suppressor gene miR-145-5p is expressed in RCC tissues, but its relationship with circRNAs is unknown. Thus, we aimed to identify differentially expressed circRNAs in RCC tissues and to explore the interaction between these circRNAs and miR-145 in the development of RCC.

methodsWe performed high-throughput sequencing and bioinformatics analyses to examine the expression pattern of circRNAs in RCC. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses were used to functionally annotate differentially expressed circRNAs. Quantitative real-time polymerase chain reaction (qRT-PCR) was used for sequence verification. Small interfering RNAs were employed to investigate the function and mechanism of circRNAs in RCC. The relationship between miR-145-5p and circRNAs was confirmed using luciferase, RNA immunoprecipitation (RIP), and biotin-coupled probe RNA pull-down assays.

resultsFifty-three circRNAs were significantly and differentially expressed in RCC compared to normal control tissue. Bioinformatic analyses indicated that two significantly upregulated circRNAs, circ-AFF2 and circ-ASAP1, had sequences corresponding to miR-145 response elements. Consistently, the luciferase reporter, RIP, and biotin-coupled probe RNA pull-down assays showed that circ-AFF2 and circ-ASAP1 may repress miR-145 by acting as sponges. circ-AFF2 and circ-ASAP1 were highly expressed in RCC patient-derived tumor samples; their overexpression correlated with poor prognosis and low miR-145 levels. Knockdown of circ-AFF2 or circ-ASAP1 in RCC cell lines inhibited proliferation, underscoring their oncogenic function. A circRNA-miRNA network was constructed for RCC using the differentially expressed circRNAs and projected miRNAs. Candidate genes were verified by RT-qPCR and western blot, indicating that circ-AFF2 and circ-ASAP1 may be connected to RCC proliferation and metastasis.

conclusioncirc-AFF2 and circ-ASAP1 were upregulated in RCC and likely promote tumor progression by sponging miR-145. Therefore, both circRNAs should be investigated further as potential diagnostic and therapeutic targets for RCC.