Transcriptomic analysis of gastric cancer cells in response to stable over-expression of circRNA derived from hnRNPU (circ-hnRNPU)

Circular RNA (circRNA) is a type of endogenous single-stranded and covalently closed non-coding RNA that plays emerging roles in tumorigenesis and aggressiveness. However, the functions and underlying mechanisms of circRNAs in regulating gastric cancer progression still remain elusive. We identify circ-hnRNPU, a circRNA derived from exons 5, 6, 7 and 8 of heterogeneous nuclear ribonucleoprotein U (hnRNPU), as a tumor suppressor of gastric cancer progression. To investigate the mechanisms underlying the functions of circ-hnRNPU, we employed the Illumina HiSeq X Ten as a discovery platform to analyze the transcriptome profiling changes of human gastric cancer MKN-45 and AGS cells in response to stable over-expression of circ-hnRNPU. The results showed that stable over-expression of circ-hnRNPU led to altered expression of 112 human mRNAs, including 64 up-regulated and 48 down-regulated genes, in both MKN-45 and AGS cells. Furthermore, we validated the RNA-seq results by real-time RT-PCR with high identity. Overall, our results provided fundamental information about the transcriptomic changes in response to circ-hnRNPU over-expression in human gastric cancer cells, and these findings will help us understand the pathogenesis of cancer progression. Total RNA of cells stably transfected with empty vector or circ-hnRNPU was extracted using the TRIzol® reagent according to the manufacturer's instructions. RNA concentration was measured using a Qubit® RNA Assay Kit with a Qubit® 2.0 Fluorometer (Life Technologies, Inc.), and integrity was assessed using the RNA Nano 6000 Assay Kit with a Bioanalyzer 2100 system (Agilent Technologies, CA). Library preparation and transcriptome sequencing on an Illumina HiSeq X Ten platform were performed by Wuhan SeqHealth Technology Co., Ltd. (Wuhan, China), and 100 bp paired-end reads were generated. HTSeq v0.6.0 was used to count the reads numbers mapped to each gene.