Li Minxia, Du Ning, Tang Ping
Department of Laboratory Medicine, The People's Hospital of Danyang, Zhenjiang City, Jiangsu Province, China.
Department of Laboratory Medicine, Lingcheng District Hospital of Traditional Chinese Medicine, Dezhou City, Shandong Province, China.
Mol Cell Biochem. 2025 Sep 10. doi: 10.1007/s11010-025-05378-w.
5-methylcytosine (m5C) methylation is a post-transcriptional modification of RNAs, and its dysregulation plays pro-tumorigenic roles in lung adenocarcinoma (LUAD). Here, this study elucidated the mechanism of action of NSUN2, a major m5C methyltransferase, on LUAD progression. mRNA expression was analyzed by quantitative PCR. Protein expression was tested by immunohistochemistry and immunoblotting. Cell proliferation was evaluated by MTT and EdU assays. Cell apoptosis was detected by flow cytometry and TUNEL staining. Transwell assays were used to examine cell invasion and migration. The influence of NSUN2 or the m5C reader YBX1 in CCT5 mRNA was analyzed by RNA immunoprecipitation assay and Actinomycin D treatment. In human LUAD, NSUN2 expression was upregulated, and high NSUN2 expression foreboded worse overall survival of LUAD patients. NSUN2 knockdown suppressed cell proliferation, diminished invasive and migratory potentials, and stimulated apoptosis in LUAD cells. NSUN2 was positively associated with CCT5 expression in LUAD and could mediate m5C modification of CCT5 mRNA. NSUN2 enhanced CCT5 mRNA stability and protein expression via two specific cysteines (C271 and C321). Moreover, YBX1 promoted CCT5 mRNA stability and protein expression. Additionally, restoration of CCT5 expression abolished NSUN2 deletion-mediated in vitro anti-proliferation, pro-apoptosis, anti-migration, and anti-invasiveness effects in LUAD cells, as well as in vivo anti-tumor growth behavior. Our study indicates that NSUN2 enhances the stability of CCT5 mRNA through a YBX1-m5C-dependent manner to drive LUAD progression. Targeting the NSUN2/CCT5 axis may be a potential approach to combat LUAD.
5-甲基胞嘧啶（m5C）甲基化是RNA的一种转录后修饰，其失调在肺腺癌（LUAD）中发挥促肿瘤作用。在此，本研究阐明了主要的m5C甲基转移酶NSUN2对LUAD进展的作用机制。通过定量PCR分析mRNA表达。通过免疫组织化学和免疫印迹检测蛋白质表达。通过MTT和EdU试验评估细胞增殖。通过流式细胞术和TUNEL染色检测细胞凋亡。采用Transwell试验检测细胞侵袭和迁移。通过RNA免疫沉淀试验和放线菌素D处理分析NSUN2或m5C阅读器YBX1对CCT5 mRNA的影响。在人类LUAD中，NSUN2表达上调，高NSUN2表达预示着LUAD患者的总体生存率更差。敲低NSUN2可抑制LUAD细胞的增殖，降低其侵袭和迁移能力，并促进细胞凋亡。在LUAD中，NSUN2与CCT5表达呈正相关，并可介导CCT5 mRNA的m5C修饰。NSUN2通过两个特定的半胱氨酸（C271和C321）增强CCT5 mRNA的稳定性和蛋白质表达。此外，YBX1促进CCT5 mRNA的稳定性和蛋白质表达。此外，恢复CCT5表达消除了NSUN2缺失介导的LUAD细胞体外抗增殖、促凋亡、抗迁移和抗侵袭作用，以及体内抗肿瘤生长行为。我们的研究表明，NSUN2通过YBX1-m5C依赖性方式增强CCT5 mRNA的稳定性，从而推动LUAD进展。靶向NSUN2/CCT5轴可能是对抗LUAD的一种潜在方法。
