Lv Jiancheng, Song Qiang, Bai Kexin, Han Jie, Yu Hao, Li Kai, Zhuang Juntao, Yang Xiao, Yang Haiwei, Lu Qiang
Department of Urology, The First Affiliated Hospital of Nanjing Medical University, No. 300 Guangzhou Road, Nanjing, 210029, China.
Cancer Cell Int. 2022 Oct 5;22(1):301. doi: 10.1186/s12935-022-02701-z.
Single-nucleotide polymorphisms (SNPs) in N6-methyladenosine (m6A) related genetic locus play significant roles in tumorigenesis and development. The expression level of many oncogenes and tumour suppressor genes changed because of m6A-associated SNPs. In addition, the relationship between m6A-SNP and bladder cancer (BCa) has not been well studied.
We screened m6A-SNPs in BCa by combining m6A-SNPs data and GWAS-SNPs data. Expression quantitative trait loci (eQTL) and differential expression gene (DEGs) analyses were performed. In ring finger protein, transmembrane 2 (RNFT2), rs3088107 (C > G) was found to have significant eQTL signals and make RNFT2 gene differentially-regulated mostly in BCa. We validated the expression level of RNFT2 in 32 pairs of BCa tissues and eight BCa cell lines by quantitative real-time PCR (qRT-PCR). Functional assays were performed to investigate the role of rs3088107 and RNFT2 in BCa in vitro.
We identified 673 m6A-SNPs, which were associated with BCa. Of these m6A-SNPs, 221 showed eQTL signals, amongst which, rs3088107 in RNFT2 showed significant eQTL signals. Results of bioinformatic analyses showed that 11 genes with m6A-SNPs had a differential expression level in BCa. RNFT2 was predicted to be significantly up-regulated in BCa. The qRT-PCR results validated that RNFT2 was highly expressed in our own BCa tissues and cell lines. High expression of RNFT2 also indicated a worse overall survival. We also revealed that rs3088107 (C > G) could inhibit the expression and m6A modification of RNFT2 by qRT-PCR, western-blot and m6A-RIP assays. Moreover, the results of functional assays indicated that RNFT2 promoted BCa cell proliferation and migration.
This research found that m6A-SNPs were associated with oncogene RNFT2 in BCa. Furthermore, m6A-SNPs showed great application potential as a new BCa diagnostic biomarker and prognostic indicator.
N6-甲基腺苷（m6A）相关基因座中的单核苷酸多态性（SNP）在肿瘤发生和发展中起重要作用。许多癌基因和肿瘤抑制基因的表达水平因m6A相关的SNP而改变。此外，m6A-SNP与膀胱癌（BCa）之间的关系尚未得到充分研究。
我们通过整合m6A-SNP数据和全基因组关联研究（GWAS）-SNP数据，在BCa中筛选m6A-SNP。进行了表达定量性状基因座（eQTL）和差异表达基因（DEG）分析。在跨膜环指蛋白2（RNFT2）中，发现rs3088107（C＞G）具有显著的eQTL信号，并且主要在BCa中使RNFT2基因受到差异调节。我们通过定量实时PCR（qRT-PCR）验证了32对BCa组织和8种BCa细胞系中RNFT2的表达水平。进行功能试验以研究rs3088107和RNFT2在体外BCa中的作用。
我们鉴定出673个与BCa相关的m6A-SNP。在这些m6A-SNP中，221个显示出eQTL信号，其中RNFT2中的rs3088107显示出显著的eQTL信号。生物信息学分析结果表明，11个具有m6A-SNP的基因在BCa中的表达水平存在差异。预测RNFT2在BCa中显著上调。qRT-PCR结果验证了RNFT2在我们自己的BCa组织和细胞系中高表达。RNFT2的高表达也表明总体生存率较差。我们还通过qRT-PCR、蛋白质免疫印迹和m6A-RIP试验揭示，rs3088107（C＞G）可抑制RNFT2的表达和m6A修饰。此外，功能试验结果表明，RNFT2促进BCa细胞增殖和迁移。
本研究发现m6A-SNP与BCa中的癌基因RNFT2相关。此外，m6A-SNP作为一种新的BCa诊断生物标志物和预后指标具有巨大的应用潜力。
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