PDF(Pubmed)
Abstract：
TGF-β1 plays a pivotal role in the metastatic cascade of malignant neoplasms. N6-methyladenosine (m6A) stands as one of the most abundant modifications on the mRNA transcriptome. However, in the metastasis of gallbladder carcinoma (GBC), the effect of TGF-β1 with mRNA m6A modification, especially the effect of mRNA translation efficiency associated with m6A modification, remains poorly elucidated. Here we demonstrated a negative correlation between FOXA1 and TGF-β1 expression in GBC. Overexpression of FOXA1 inhibited TGF-β1-induced migration and epithelial-mesenchymal transition (EMT) in GBC cells. Mechanistically, we confirmed that TGF-β1 suppressed the translation efficiency of FOXA1 mRNA through polysome profiling analysis. Importantly, both in vivo and in vitro experiments showed that TGF-β1 promoted m6A modification on the coding sequence (CDS) region of FOXA1 mRNA, which was responsible for the inhibition of FOXA1 mRNA translation by TGF-β1. We demonstrated through MeRIP and RIP assays, dual-luciferase reporter assays and site-directed mutagenesis that ALKBH5 promoted FOXA1 protein expression by inhibiting m6A modification on the CDS region of FOXA1 mRNA. Moreover, TGF-β1 inhibited the binding capacity of ALKBH5 to the FOXA1 CDS region. Lastly, our study confirmed that overexpression of FOXA1 suppressed lung metastasis and EMT in a nude mice lung metastasis model. In summary, our research findings underscore the role of TGF-β1 in regulating TGF-β1/FOXA1-induced GBC EMT and metastasis by inhibiting FOXA1 translation efficiency through m6A modification.
摘要：
TGF-β1在恶性肿瘤的转移级联中起关键作用。N6-甲基腺苷（m6A）是mRNA转录组上最丰富的修饰之一。然而,在胆囊癌（GBC）的转移中，TGF-β1对mRNAm6A修饰的影响，特别是与m6A修饰相关的mRNA翻译效率的影响，仍未阐明。在这里，我们证明了FOXA1和TGF-β1在GBC中的表达之间呈负相关。FOXA1过表达抑制TGF-β1诱导的GBC细胞迁移和上皮-间质转化(EMT)。机械上,我们通过多聚体谱分析证实TGF-β1抑制FOXA1mRNA的翻译效率.重要的是,体内和体外实验均表明TGF-β1促进FOXA1mRNA编码序列(CDS)区的m6A修饰,其负责通过TGF-β1抑制FOXA1mRNA翻译。我们通过MeRIP和RIP试验证明，ALKBH5通过抑制FOXA1mRNACDS区的m6A修饰促进FOXA1蛋白表达的双荧光素酶报告基因测定和定点诱变。此外，TGF-β1抑制ALKBH5与FOXA1CDS区的结合能力。最后,我们的研究证实FOXA1的过表达抑制了裸鼠肺转移模型中的肺转移和EMT。总之,我们的研究结果强调了TGF-β1通过m6A修饰抑制FOXA1翻译效率,在调节TGF-β1/FOXA1诱导的GBCEMT和转移中的作用.
