Two groups of control and RANBP1-knockdown HCT116 cells

Colorectal cancer (CRC) is among the top five most common malignant tumors worldwide and has a high mortality rate. Identification of the mechanism of CRC and potential therapeutic targets is critical for improving survival. In the present study, we observed high expression of RAN binding protein 1 (RANBP1) in CRC tissues. Upregulated RANBP1 expression was strongly associated with TNM stages and was an independent risk factor for poor prognosis. In vitro and in vivo functional experiments demonstrated that RANBP1 promoted the proliferation and invasion of CRC cells, affected the cell cycle, and inhibited the apoptosis of CRC cells. Low RANBP1 expression reduced the expression levels of hsa-miR-18a, hsa-miR-183, and hsa-miR-106 microRNAs (miRNAs) by inhibiting the nucleoplasmic transport of precursor miRNAs (pre-miRNAs), thereby promoting the accumulation of the latter in the nucleus and reducing the expression of mature miRNAs. Further experiments and bioinformatic analyses demonstrated that RANBP1 promoted the expression of YAP by regulating miRNAs and the Hippo pathway. We also found that YAP acted as a transcriptional cofactor to activate RANBP1 transcription in combination with TEAD4 transcription factor. Thus, RANBP1 further promoted the progression of CRC by forming a positive feedback loop with YAP. Our results revealed the biological role and mechanism of RANBP1 in CRC for the first time, suggesting that RANBP1 can be used as a diagnostic molecule and a potential therapeutic target in CRC. Overall design: Total RNA, including small RNAs from two groups of control and RANBP1-knockdown HCT116 cells, were extracted using the Total RNA Purification Kit from Norgen Biotek Corp (Thorold, Canada). Approximately 1 µg total RNA was used to prepare a small RNA library according to the protocol of the TruSeq Small RNA Sample Prep Kit (Illumina, San Diego, CA). Then, single-end sequencing (36 bp or 50 bp) was performed on the Illumina HiSeq 2500 System (LC Sciences, Houston, TX) according to the manufacturer's recommended protocol. Raw reads were processed with ACGT101-miR (LC Sciences, Houston, Texas, USA) to remove adapter dimers, junk, low complexity reads, common RNA families (rRNA, tRNA, snRNA, and snoRNA) and repeats. Subsequently, unique sequences containing 18–26 bases were mapped to specific species precursors in miRBase 22.0 using a BLAST search to identify known miRNAs and novel miRNA-3p- and miRNA-5p-derived miRNAs. The mapped sequences were identified as known miRNAs. The remaining sequences were mapped to other selected species precursors in miRBase 22.0 by BLAST search. Differential expression of miRNAs as a function of normalized deep sequence counting was analyzed through the selective use of Student's t-test. The significance threshold was set at <0.05.