Ding Chengming, Tao Guangwei, Chen Guodong, Xie Yi, Yang Chunfen, Qi Shuo, Hou Jiafeng, Jiang Xinmiao, Deng Xin, Liao Wenyan
The First Affiliated Hospital, Department of Hepatopancreatobiliary Surgery, Hengyang Medical School, University of South China, Hengyang, Hunan 421001, China.
The First Affiliated Hospital, Department of Gynaecology and Obstetrics, Hengyang Medical School, University of South China, Hengyang, Hunan 421001, China.
Mol Immunol. 2024 Dec;176:73-83. doi: 10.1016/j.molimm.2024.11.002. Epub 2024 Nov 24.
Phosphoribosylformylglycinamidine synthase (PFAS) is a critical enzyme in de novo synthesis of purine. Innate immunity recognizes tumor derived damage-associated molecular patterns (DAMPs) and initiates the anti-tumor adaptive responses. While the function of PFAS catalyzed de novo synthesis of purine is well proved, its effect on innate immune evasion in cancer is unclear and needs to be further explored. The purpose of this study was to investigate the specific mechanisms by which PFAS inhibits RIG-I receptor (RLR) -mediated NF-κB axis in CRC.
quantitative real-time PCR (qRT-PCR), Immunohistochemical (IHC) staining and western blotting were conducted to study the expression of PFAS in CRC tissues. Survival analysis, COX regression analysis and receiver operating characteristic (ROC) curve analysis were respectively conducted to assess correlation between the PFAS expression and clinicopathological characteristics, investigate the percent survival based on PFAS level in different clinical CRC groups, identify factors influencing the prognosis of CRC, and illustrate the diagnostic ability of PFAS in CRC patients. Furthermore, the CCK8 and transwell assays were carried out to study CRC cell function affected by PFAS. Mechanistically, plaque assay was used to assess the regulation of PFAS on innate immune signalling. The inhibition of PFAS on RIG-I-mediated innate immune signalling was further investigated by qRT-PCR and reporter assays in thepresence of lentiviral-mediated PFAS stably knocking down and stably overexpressing. Lastly, the interaction between PFAS and RIG-I was verified by co-immunoprecipitation assay.
The expression of PFAS in CRC tissue was higher than in adjacent normal colorectal tissue. The level of PFAS expression was significantly associated with stage-AJCC, regional lymph nodes metastasis and recurrence in CRC. Low expression of gene PFAS caused better survival than high expression in CRC patients. PFAS could be considered as an independent prognostic risk factor of CRC. PFAS promote cell proliferation and invasion of CRC cell lines. According to ROC curve analysis, PFAS could be used as a diagnostic biomarker in CRC. Mechanistically, PFAS inhibit interferon-β (IFN-β) gene and interferon-stimulated gene 56 (ISG56) expression. Furthermore, we confirmed that PFAS target RIG-I to inhibit RIG-I-mediated innate immune signalling.
磷酸核糖甲酰甘氨脒合成酶（PFAS）是嘌呤从头合成中的关键酶。固有免疫识别肿瘤来源的损伤相关分子模式（DAMPs）并启动抗肿瘤适应性反应。虽然PFAS催化嘌呤从头合成的功能已得到充分证实，但其在癌症中对固有免疫逃逸的影响尚不清楚，需要进一步探索。本研究的目的是探讨PFAS抑制结直肠癌中RIG-I受体（RLR）介导的NF-κB轴的具体机制。
采用定量实时PCR（qRT-PCR）、免疫组织化学（IHC）染色和蛋白质印迹法研究PFAS在结直肠癌组织中的表达。分别进行生存分析、COX回归分析和受试者工作特征（ROC）曲线分析，以评估PFAS表达与临床病理特征之间的相关性，基于不同临床结直肠癌组中PFAS水平研究生存率，确定影响结直肠癌预后的因素，并阐明PFAS在结直肠癌患者中的诊断能力。此外，进行CCK8和Transwell实验以研究PFAS对结直肠癌细胞功能的影响。从机制上讲，采用噬斑实验评估PFAS对固有免疫信号的调节作用。在慢病毒介导的PFAS稳定敲低和稳定过表达的情况下，通过qRT-PCR和报告基因实验进一步研究PFAS对RIG-I介导的固有免疫信号的抑制作用。最后，通过免疫共沉淀实验验证PFAS与RIG-I之间的相互作用。
PFAS在结直肠癌组织中的表达高于相邻正常结直肠组织。PFAS的表达水平与结直肠癌的AJCC分期、区域淋巴结转移和复发显著相关。基因PFAS低表达的结直肠癌患者生存率高于高表达患者。PFAS可被视为结直肠癌的独立预后危险因素。PFAS促进结直肠癌细胞系的细胞增殖和侵袭。根据ROC曲线分析，PFAS可作为结直肠癌的诊断生物标志物。从机制上讲，PFAS抑制干扰素-β（IFN-β）基因和干扰素刺激基因56（ISG56）的表达。此外，我们证实PFAS靶向RIG-I以抑制RIG-I介导的固有免疫信号。
