Wang Changli, Liu Wanting, Yang Sheng, Wang Tianyi, Wang Bing
Institute of Biochemistry and Molecular Biology, College of Life and Health Sciences, Northeastern University, Shenyang 110819, China.
Int J Mol Sci. 2025 Jun 7;26(12):5483. doi: 10.3390/ijms26125483.
This study explores the regulatory function of BAP31 on exosomal miRNA and its impact on the EMT in CRC. Exosomes from BAP31-OE cells promoted recipient cell migration and triggered the EMT, as indicated by decreased E-cadherin and increased N-cadherin and Vimentin levels. By contrast, exosomes derived from shBAP31 cells were observed to inhibit cell migration and revert EMT markers. The administration of shBAP31 exosomes significantly inhibited tumor growth in vivo. miRNA profiling revealed 76 differentially expressed miRNAs in BAP31-OE exosomes. Six miRNA candidates associated with the EMT were identified in the GEO database, miR-423-3p was identified as a key mediator, the candidates from shBAP31 exosomes exhibited the opposite effect. EMT promotion by miR-423-3p was further evidenced by EMT marker expression, enhanced migratory capacity, and accelerated tumor growth. Sixteen potential target genes were identified through bioinformatics analysis. Bim exhibited significant downregulation by the miR-423-3p mimic. Luciferase reporter assays verified the direct interaction between miR-423-3p and the 3'UTR of Bim. Silencing Bim negated the effects of miR-423-3p. It was also revealed that BAP31 does not influence the total exosomal miRNA content but selectively regulates miR-423-3p, which contains an EXOmotif enriched in BAP31-OE exosomes. Mechanistic studies revealed that BAP31 enhances the expression of the RNA export adaptor Alyref, as validated by qRT-PCR and Western blot analyses. RNA immunoprecipitation assays verified that Alyref binds to miR-423-3p in BAP31-OE cells. Our results reveal that BAP31 facilitates the sorting of exosomal miR-423-3p via Alyref, thereby promoting EMT in CRC through the miR-423-3p/Bim signaling axis. This indicates that BAP31 could be a viable therapeutic target for managing the EMT in CRC.
本研究探讨了BAP31对外泌体miRNA的调控作用及其对结直肠癌上皮-间质转化（EMT）的影响。BAP31过表达（OE）细胞来源的外泌体促进受体细胞迁移并触发EMT，表现为E-钙黏蛋白水平降低，N-钙黏蛋白和波形蛋白水平升高。相比之下，观察到短发夹RNA干扰BAP31（shBAP31）细胞来源的外泌体抑制细胞迁移并使EMT标志物逆转。给予shBAP31外泌体可显著抑制体内肿瘤生长。miRNA谱分析显示BAP31-OE外泌体中有76种差异表达的miRNA。在基因表达 omnibus 数据库（GEO数据库）中鉴定出6种与EMT相关的miRNA候选物，其中miR-423-3p被确定为关键介质，shBAP31外泌体中的候选物表现出相反的作用。miR-423-3p对EMT的促进作用通过EMT标志物表达、增强的迁移能力和加速的肿瘤生长得到进一步证实。通过生物信息学分析鉴定出16个潜在靶基因。miR-423-3p模拟物使Bim显著下调。荧光素酶报告基因检测验证了miR-423-3p与Bim的3'非翻译区（3'UTR）之间的直接相互作用。沉默Bim可消除miR-423-3p的作用。研究还表明，BAP31不影响外泌体miRNA的总含量，但选择性地调节miR-423-3p，其包含在BAP31-OE外泌体中富集的EXOmotif。机制研究表明，BAP31增强了RNA输出衔接蛋白Alyref的表达，这通过定量逆转录聚合酶链反应（qRT-PCR）和蛋白质免疫印迹分析得到验证。RNA免疫沉淀试验证实Alyref在BAP31-OE细胞中与miR-423-3p结合。我们的结果表明，BAP31通过Alyref促进外泌体miR-423-3p的分选，从而通过miR-423-3p/Bim信号轴促进结直肠癌中的EMT。这表明BAP31可能是控制结直肠癌中EMT的一个可行治疗靶点。
