放射抗性是晚期宫颈癌（CC）死亡的主要原因。 RNA 修饰失调最近已成为辐射和耐药性的调节机制。我们的目的是探讨5-甲基胞嘧啶（m5C）在宫颈癌放射敏感性中的生物学功能和临床意义。通过液相色谱-串联质谱法对放疗耐药和敏感CC标本中RNA修饰的丰度进行定量。通过RNA测序筛选与CC放射敏感性相关的必需RNA修饰相关基因。 NSUN6 对放射敏感性的影响在 CC 细胞系、细胞源性异种移植物 (CDX) 和 3D 生物打印患者源性类器官 (PDO) 中得到验证。通过综合m5C测序、mRNA测序和RNA免疫沉淀研究NSUN6调节CC放射敏感性的机制。我们发现在耐药CC样本中m5C修饰丰度较高，NSUN6是与放射敏感性相关的重要m5C调节基因。 NSUN6 过度表达在临床上与宫颈癌的放射抗性和不良预后相关。从功能上讲，较高的 NSUN6 表达与宫颈癌 3D PDO 模型中的放射抗性相关。此外，沉默 NSUN6 会增加 CC 在体内和体外的放射敏感性。从机制上讲，NDRG1 是通过集成 m5C-seq、mRNA-seq 和功能验证鉴定的 NSUN6 下游靶基因之一。 NSUN6促进NDRG1 mRNA的m5C修饰，并且m5C阅读器ALYREF与m5C标记的NDRG1 mRNA明确结合并增强NDRG1 mRNA稳定性。 NDRG1 过表达促进同源重组介导的 DNA 修复，进而导致宫颈癌的放射抵抗。异常的 m5C 高甲基化和 NSUN6 过表达导致宫颈癌对放射治疗的抵抗。 NSUN6 表达升高通过激活 NSUN6/ALYREF-m5C-NDRG1 通路促进宫颈癌的放射抗性。 NSUN6 在宫颈癌中的低表达表明对放疗的敏感性和更好的预后。© 2024。作者。

Radioresistance is the leading cause of death in advanced cervical cancer (CC). Dysregulation of RNA modification has recently emerged as a regulatory mechanism in radiation and drug resistance. We aimed to explore the biological function and clinical significance of 5-methylcytosine (m5C) in cervical cancer radiosensitivity.The abundance of RNA modification in radiotherapy-resistant and sensitive CC specimens was quantified by liquid chromatography-tandem mass spectrometry. The essential RNA modification-related genes involved in CC radiosensitivity were screened via RNA sequencing. The effect of NSUN6 on radiosensitivity was verified in CC cell lines, cell-derived xenograft (CDX), and 3D bioprinted patient-derived organoid (PDO). The mechanisms of NSUN6 in regulating CC radiosensitivity were investigated by integrative m5C sequencing, mRNA sequencing, and RNA immunoprecipitation.We found a higher abundance of m5C modification in resistant CC samples, and NSUN6 was the essential m5C-regulating gene concerning radiosensitivity. NSUN6 overexpression was clinically correlated with radioresistance and poor prognosis in cervical cancer. Functionally, higher NSUN6 expression was associated with radioresistance in the 3D PDO model of cervical cancer. Moreover, silencing NSUN6 increased CC radiosensitivity in vivo and in vitro. Mechanistically, NDRG1 was one of the downstream target genes of NSUN6 identified by integrated m5C-seq, mRNA-seq, and functional validation. NSUN6 promoted the m5C modification of NDRG1 mRNA, and the m5C reader ALYREF bound explicitly to the m5C-labeled NDRG1 mRNA and enhanced NDRG1 mRNA stability. NDRG1 overexpression promoted homologous recombination-mediated DNA repair, which in turn led to radioresistance in cervical cancer.Aberrant m5C hypermethylation and NSUN6 overexpression drive resistance to radiotherapy in cervical cancer. Elevated NSUN6 expression promotes radioresistance in cervical cancer by activating the NSUN6/ALYREF-m5C-NDRG1 pathway. The low expression of NSUN6 in cervical cancer indicates sensitivity to radiotherapy and a better prognosis.© 2024. The Author(s).