Ying Shuai, Wu Ningxia, Ruan Yuting, Ge Wen, Ma Pei, Xu Tongpeng, Shu Yongqian, Wang Yingwei, Qiu Wen, Zhao Chenhui
Department of Immunology, Nanjing Medical University, Nanjing, 211166, China.
Department of Laboratory Medicine, Wuxi People's Hospital of Nanjing Medical University, Wuxi, 214023, China.
BMC Cancer. 2025 Jan 14;25(1):81. doi: 10.1186/s12885-025-13473-w.
Non-small cell lung cancer (NSCLC) is a disease related to inflammation. Proinflammatory cytokines such as interleukin 17 (IL-17) can induce cancer cell proliferation, metastasis and immune escape. Although NSCLC immune escape is partly due to the interaction between PD-1 and PD-L1 and PD-L1 expression can be upregulated in cancer cells upon stimulation with IL-17, the underlying mechanism of IL-17-triggered PD-L1 gene transcription in NSCLC cells remains elusive.
RT‒PCR, real-time PCR, and IB were used to assess the levels of PD-L1, MEF2C, and TRIM31 in NSCLC tissues as well as in IL-17-stimulated H1299 or PC9 cells. Bioinformatics analysis, luciferase assays, and ChIP were utilized to investigate the transcriptional mechanism of the PD-L1 gene. Co-IP/IB was used to examine the interaction between MEF2C and PD-L1, including MEF2C ubiquitination. IHC staining was carried out to analyse the expression of IL-17RA, MEF2C, TRIM31, and PD-L1 in NSCLC tissue arrays. The corresponding plasmids were constructed and identified. An isograft model was used to verify the findings in vitro.
PD-L1, MEF2C and TRIM31 expression levels were increased in NSCLC tissues and NSCLC cells exposed to IL-17. Mechanistically, MEF2C could bind to the - 778 to -475 nt and - 336 to -97 nt regions of the PD-L1 promoter. TRIM31 could mediate MEF2C K63-linked polyubiquitination at Lys 25, increasing MEF2C recruitment to the PD-L1 promoter and PD-L1 gene transcription. MEF2C, TRIM31 or PD-L1 gene silencing effectively suppressed MEF2C K63-linked polyubiquitination, PD-L1 induction and NSCLC growth in mice inoculated with Lewis lung cancer (LLC) cells transfected with the corresponding shRNA and treated with IL-17.
IL-17 induces PD-L1 gene transcription in NSCLC cells through TRIM31-dependent MEF2C K63-linked polyubiquitination.
非小细胞肺癌（NSCLC）是一种与炎症相关的疾病。白细胞介素17（IL-17）等促炎细胞因子可诱导癌细胞增殖、转移和免疫逃逸。尽管NSCLC免疫逃逸部分归因于PD-1与PD-L1之间的相互作用，且IL-17刺激后癌细胞中PD-L1表达可上调，但IL-17触发NSCLC细胞中PD-L1基因转录的潜在机制仍不清楚。
采用逆转录聚合酶链反应（RT-PCR）、实时荧光定量PCR和免疫印迹法（IB）评估NSCLC组织以及IL-17刺激的H1299或PC9细胞中PD-L1、MEF2C和TRIM31的水平。利用生物信息学分析、荧光素酶报告基因检测和染色质免疫沉淀法（ChIP）研究PD-L1基因的转录机制。采用免疫共沉淀/免疫印迹法（Co-IP/IB）检测MEF2C与PD-L1之间的相互作用，包括MEF2C的泛素化。进行免疫组化染色分析NSCLC组织芯片中IL-17RA、MEF2C、TRIM31和PD-L1的表达。构建并鉴定相应质粒。采用同基因移植模型在体外验证研究结果。
NSCLC组织和暴露于IL-17的NSCLC细胞中，PD-L1、MEF2C和TRIM31表达水平升高。机制上，MEF2C可与PD-L1启动子的-778至-475 nt和-336至-97 nt区域结合。TRIM31可介导MEF2C在赖氨酸25位点发生K63连接的多聚泛素化，增加MEF2C对PD-L1启动子的募集及PD-L1基因转录。MEF2C、TRIM31或PD-L1基因沉默可有效抑制接种用相应短发夹RNA（shRNA）转染并经IL-17处理的Lewis肺癌（LLC）细胞的小鼠中MEF2C的K63连接多聚泛素化、PD-L1诱导及NSCLC生长。
IL-17通过TRIM31依赖的MEF2C K63连接多聚泛素化诱导NSCLC细胞中PD-L1基因转录。
