Department of Thoracic Surgery, Third Hospital of Shanxi Medical University, Shanxi Bethune Hospital, Shanxi Academy of Medical Sciences, Tongji Shanxi Hospital, Taiyuan, China.
Department of Biochemistry and Molecular Biology, Shanxi Medical University, Taiyuan, China.
RNA Biol. 2023 Jan;20(1):359-367. doi: 10.1080/15476286.2023.2223939.
Often, alternative splicing is used by cancer cells to produce or increase proteins that promote growth and survival through alternative splicing. Although RNA-binding proteins are known to regulate alternative splicing events associated with tumorigenesis, their role in oesophageal cancer (EC) has rarely been explored.
We analysed the expression pattern of several relatively well characterized splicing regulators on 183 samples from TCGA cohort of oesophageal cancer; the effectiveness of the knockdown of SRSF2 was subsequently verified by immunoblotting; we measured the ability of cells treated with lenti-sh-SRSF2/lenti-sh2-SRSF2 to invade through an extracellular matrix coating by transwell invasion assay; using RNA-seq data to identify its potential target genes; we performed qRT-PCR to detect the changes of exon 2 usage in lenti-sh-SRSF2 transduced KYSE30 cells to determine the possible effect of SRSF2 on splicing regulation of IRF3; RNA Electrophoretic mobility shift assay (RNA-EMSA) was performed by the incubation of purified SRSF2 protein and biotinylated RNA probes; we performed luciferase assay to confirm the effect of SRSF2 on IFN1 promoter activity.
We found upregulation of SRSF2 is correlated with the development of EC; Knock-down of SRSF2 inhibits EC cell proliferation, migration, and invasion; SRSF2 regulates the splicing pattern of IRF3 in EC cells; SRSF2 interacts with exon 2 of IRF3 to regulate its exclusion; SRSF2 inhibits the transcription of IFN1 in EC cells.
This study identified a novel regulatory axis involved in EC from the various aspects of splicing regulation.
癌细胞经常通过选择性剪接产生或增加促进生长和存活的蛋白质。虽然已知 RNA 结合蛋白可调节与肿瘤发生相关的选择性剪接事件，但它们在食管癌 (EC) 中的作用很少被探索。
我们分析了 TCGA 队列中 183 个食管癌样本中几种相对特征明确的剪接调节剂的表达模式；随后通过免疫印迹验证了 SRSF2 敲低的有效性；我们通过 Transwell 侵袭测定测量了用 lenti-sh-SRSF2/lenti-sh2-SRSF2 处理的细胞穿过细胞外基质涂层的侵袭能力；使用 RNA-seq 数据来识别其潜在的靶基因；我们进行 qRT-PCR 检测 lenti-sh-SRSF2 转导的 KYSE30 细胞中外显子 2 使用率的变化，以确定 SRSF2 对 IRF3 剪接调节的可能影响；通过孵育纯化的 SRSF2 蛋白和生物素化 RNA 探针进行 RNA 电泳迁移率变动分析 (RNA-EMSA)；我们进行荧光素酶测定以确认 SRSF2 对 IFN1 启动子活性的影响。
我们发现 SRSF2 的上调与 EC 的发生有关；SRSF2 的敲低抑制 EC 细胞的增殖、迁移和侵袭；SRSF2 调节 EC 细胞中 IRF3 的剪接模式；SRSF2 与 IRF3 的外显子 2 相互作用以调节其排除；SRSF2 抑制 EC 细胞中 IFN1 的转录。
本研究从剪接调节的各个方面鉴定了涉及 EC 的新的调节轴。
