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中华肺部疾病杂志(电子版) ›› 2025, Vol. 18 ›› Issue (02) : 273 -278. doi: 10.3877/cma.j.issn.1674-6902.2025.02.013

论著

circUBAP2L 通过hsa-let-7c/GAD1 轴调控肺腺癌1-甲基组氨酸和二甲基甘氨酸的机制分析
夏祯祎1, 刘懿1, 丁俞成1, 袁宏霞2, 李晓燕2, 陈田2,()   
  1. 1. 404000 重庆,重庆大学附属三峡医院胸外科
    2. 404000 重庆,重庆大学附属三峡医院健康管理中心
  • 收稿日期:2025-02-13 出版日期:2025-04-25
  • 通信作者: 陈田
  • 基金资助:
    2021重庆市自然科学基金面上项目(cstc2021jcyj-msxmX0996)2023重庆市万州区科卫联合医学科研项目(wzstc-kw2023010)

Regulation of 1-methylhistidine and dimethylglycine in lung adenocarcinoma by circUBAP2L via hsalet-7c/GAD1 axis

Zhenyi Xia1, Yi Liu1, Yucheng Ding1, Hongxia Yuan2, Xiaoyan Li2, Tian Chen2,()   

  1. 1. Department of Thoracic Surgery,Chongqing University Three Gorges Hospital,Chongqing 404000,China
    2. Health Management Center,Chongqing University Three Gorges Hospital,Chongqing 404000,China
  • Received:2025-02-13 Published:2025-04-25
  • Corresponding author: Tian Chen
引用本文:

夏祯祎, 刘懿, 丁俞成, 袁宏霞, 李晓燕, 陈田. circUBAP2L 通过hsa-let-7c/GAD1 轴调控肺腺癌1-甲基组氨酸和二甲基甘氨酸的机制分析[J/OL]. 中华肺部疾病杂志(电子版), 2025, 18(02): 273-278.

Zhenyi Xia, Yi Liu, Yucheng Ding, Hongxia Yuan, Xiaoyan Li, Tian Chen. Regulation of 1-methylhistidine and dimethylglycine in lung adenocarcinoma by circUBAP2L via hsalet-7c/GAD1 axis[J/OL]. Chinese Journal of Lung Diseases(Electronic Edition), 2025, 18(02): 273-278.

目的

分析环状泛素相关蛋白样因子2(circular ubiquitin associated protein 2-like,circUBAP2L)、hsa-let-7c 及靶基因谷氨酸脱羧酶1(glutamatedecarboxylase1,GAD1)在肺腺癌中的作用。

方法

采用small-RNA 测序发现hsa-let-7c 差异表达,通过生物信息学、实时聚合酶链反应(polymerase chain reaction,PCR)分析hsa-let-7c 上游基因circUBAP2L、下游靶基因GAD1 的差异表达。 采用双荧光素酶报告基因实验印证circUBAP2L 与hsa-let-7c、hsa-let-7c 与GAD1 的调控关系。 通过慢病毒构建A549 肺癌细胞GAD1 过表达模型,转录组学RNA 测序(RNA sequencing,RNA-seq)和代谢组学液相色谱-质谱联用(liquid chromatography-mass spectroscopy,LC-MS)分析GAD1 的差异基因及代谢物,通过多组学联合分析GAD1 对肺腺癌细胞的氨基酸代谢影响。

结果

实时PCR 实验印证circUBAP2L 在肺腺癌中表达上调(P<0.01);TCGA 和GTEx 数据库分析结果显示,肺腺癌组织中hsa-let-7c 表达降低(P<0.0001),Small-RNA 测序结果印证hsa-let-7c 在肺腺癌中表达降低(P<0.01);转录组测序显示与正常组织相比,GAD1 在肺腺癌组织中表达升高(P=0.032)。 双荧光素酶报告基因发现circUBAP2L 序列上位点(CTACCTC-CGACAACCTATACCT)与hsa-let-7c 结合,代谢组学(LC-MS)hsa-let-7c 启动子序列上GAD1 基因位点(AACAUGU)与GAD1 结合。 转录组学和代谢组学分析结果显示,肺腺癌细胞GAD1 过表达增加1-甲基组氨酸(log2FC=2.37,P=0.00023)、L-赤式-4-羟基谷氨酸(log2FC=2.02,P=0.00055)、二甲基甘氨酸(log2FC=2.45,P=0.00129)水平,影响甘氨酸、丝氨酸和苏氨酸代谢通路(P=0.00059)及癌症通路(P=0.00541)。

结论

circUBAP2L 通过hsa-let-7c/GAD1 轴调控肺腺癌细胞中1-甲基组氨酸和二甲基甘氨酸代谢。

Objective

To investigate the roles of circular ubiquitin-associated protein 2-like(circUBAP2L),hsa-let-7c,and its target gene glutamate decarboxylase 1 (GAD1) in lung adenocarcinoma.

Methods

Small-RNA sequencing was used to identify differential expression of hsa-let-7c. Bioinformatics analysis and real-time polymerase chain reaction (PCR) were used to assess differential expression of circUBAP2L (the upstream gene of hsa-let-7c) and GAD1 (the downstream target gene). Dual-luciferase reporter assays were conducted to validate the regulatory relationships between circUBAP2L and hsa-let-7c,as well as hsa-let-7c and GAD1. A lentivirus was used to construct a GAD1 overexpression model in A549 lung cancer cells. Transcriptomic RNA sequencing (RNA-seq) and metabolomic liquid chromatography-mass spectrometry (LC-MS) were performed to analyze GAD1-associated differential genes and metabolites. Multiomics integration was applied to explore the impact of GAD1 on amino acid metabolism in lung adenocarcinoma cells.

Results

Real-time PCR confirmed upregulated expression of circUBAP2L in lung adenocarcinoma (P<0.01). Analysis of TCGA and GTEx databases revealed reduced expression of hsa-let-7c in lung adenocarcinoma tissues (P <0.0001),which was further validated by small-RNA sequencing (P <0.01).Transcriptomic sequencing showed elevated GAD1 expression in lung adenocarcinoma compared to normal tissues ( P = 0. 032 ). Dual-luciferase reporter assays identified a binding site ( CTACCTCCGACAACCTATACCT) between circUBAP2L and hsa-let-7c,while LC-MS metabolomics revealed that the GAD1 gene locus (AACAUGU) on the hsa-let-7c promoter binds to GAD1. Transcriptomic and metabolomic analyses demonstrated that GAD1 overexpression in lung adenocarcinoma cells increased levels of 1-methylhistidine (log2FC=2.37,P=0.00023),L-erythro-4-hydroxyglutamate (log2FC=2.02,P=0.00055),and dimethylglycine (log2FC = 2.45,P = 0.00129),affecting glycine,serine,and threonine metabolism pathways (P=0.00059) and cancer-related pathways (P=0.00541).

Conclusion

circUBAP2L regulates 1-methylhistidine and dimethylglycine metabolism in lung adenocarcinoma cells through the hsa-let-7c/GAD1 axis.

图1 肺腺癌中GAD1、hsa-let-7c、circUBAP2L 差异表达。 图A 为GAD1 的上游miRNA 交集;图B 为hsa-let-7c 在肺腺癌组织中的表达;图C 为hsa-let-7c 在肺腺癌中的预后;图D 为circUBAP2L 与hsa-let-7c 预测结合位点
图2 GAD1 过表达肺腺癌细胞转录组中甘氨酸、丝氨酸和苏氨酸代谢通路及癌症途径精氨酸和脯氨酸代谢通路。 图A 为富集分析图为甘氨酸、丝氨酸和苏氨酸代谢通路(HSA00260); 图 B 为富集分析图: 癌症途径通路(HSA05200);图C 为富集分析图为精氨酸和脯氨酸代谢通路(HSA00330)
图3 GAD1 过表达调控肺腺癌细胞1-甲基组氨酸、L-赤式-4-羟基谷氨酸、二甲基甘氨酸水平。 图A 为GAD1 过表达肺腺癌细胞中1-甲基组氨酸水平(n=4);图B 为GAD1 过表达肺腺癌细胞中L-赤式-4-羟基谷氨酸水平(n=4);图C 为GAD1 过表达肺腺癌细胞中二甲基甘氨酸水平(n=4)
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