下调丝氨酸/精氨酸富集剪接因子6通过调控髓系分化因子88选择性剪接抑制巨噬细胞炎症反应  

Downregulation of SRSF6 Inhibits the Macrophage Inflammatory Response by Regulating the Alternative Splicing of MyD88

作　　者：付育 张璐瑶 程博 石炜业 李霓 王英泽[1] FU Yu;ZHANG Lu-Yao;CHENG Bo;SHI Wei-Ye;LI Ni;WANG Ying-Ze(College of Food Science and Biology,Hebei University of Science and Technology,Shijiazhuang 050018,Hebei,China;State Key Laboratory of Bioactive Substance and Function of Natural Medicines,Institute of Materia Medica,Chinese Academy of Medical Sciences&Peking Union Medical College,Beijing 100050,China)

机构地区：[1]河北科技大学食品与生物学院生命科学系,石家庄050018 [2]中国医学科学院&北京协和医学院药物研究所,天然药物活性物质与功能国家重点实验室,北京100050

出　　处：《中国生物化学与分子生物学报》2024年第11期1563-1573,共11页Chinese Journal of Biochemistry and Molecular Biology

基　　金：河北省自然科学基金项目(No.H2020208002);河北省高等学校科学技术研究项目(No.ZD2022011)资助。

摘　　要：丝氨酸/精氨酸富集剪接因子6(serine/arginine-rich splicing factor 6,SRSF6)属于SR蛋白家族,主要在RNA剪接中发挥调控作用。SRSF6表达或功能失调能够导致部分基因选择性剪接异常,进而引发肿瘤、糖尿病和胸膜纤维化等炎症性疾病发生发展,但是SRSF6在炎症中的作用尚未阐明。本研究采用脂多糖(lipopolysaccharides,LPS)诱导小鼠巨噬细胞RAW264.7建立炎症反应体系,探究SRSF6在炎症中的表达及作用,初步分析SRSF6在炎症中的靶点和机制。研究发现,LPS刺激肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)、环氧合酶-2(cyclooxygenase-2,COX-2)等炎症相关因子表达增加,同时,SRSF6 mRNA和蛋白质表达水平均随LPS刺激时间延长而显著增加(P<0.05)。进一步探究SRSF6表达对炎症因子的影响,上调表达SRSF6促进LPS诱导的炎症因子表达增加(P<0.05),而下调SRSF6则抑制炎症因子表达(P<0.05),并且核因子-κB(nuclear factor,NF-κB)和丝裂原活化蛋白激酶(mitogen-activated protein kinase,MAPK)信号通路的激活受到SRSF6下调的抑制(P<0.05),提示SRSF6在巨噬细胞中参与调控炎症反应。髓系分化因子88(myeloid differentiation factor 88,MyD88)是TLR4信号通路中的关键接头分子,MyD 88 mRNA剪接异构体MyD88-L和MyD88-S在炎症反应中分别发挥促炎和抑炎的相反功能。RNA结合蛋白数据库分析和RNA结合蛋白免疫沉淀实验发现,SRSF6蛋白结合MyD 88 mRNA;剪接分析结果显示,下调SRSF6促进抑炎型剪接异构体MyD88-S mRNA表达增多(P<0.01),并且敲低MyD88-S能够恢复SRSF6下调所抑制的炎症因子表达。以上研究结果发现,SRSF6在巨噬细胞中通过调控MyD 88选择性剪接,从而影响炎症信号通路激活及炎症因子表达,这为深入解析SRSF6在炎症性疾病中的作用奠定了基础。Serine/arginine-rich splicing factor 6(SRSF6)is a member of the serine and arginine-rich(SR)protein family,and it plays a crucial regulatory role in RNA splicing.Dysregulation of SRSF6 expression or function can lead to aberrant alternative splicing of certain genes,and contribute to the development and progression of inflammatory diseases,including tumors,diabetes,and pleural fibrosis.However,the role of SRSF6 in inflammation remains unclear.In this study,we found that the expression of inflammatory factors,including tumor necrosis factor-α(TNF-α)and cyclooxygenase-2(COX-2),was induced by lipopolysaccharides(LPS).Concurrently,both the levels of SRSF6 mRNA and protein expression significantly increased with prolonged LPS stimulation(P<0.05).Furthermore,we investigated the change of SRSF6 expression on the expression of inflammatory factors.The results showed that upregulation of SRSF6 enhanced the expression of LPS-induced inflammatory factors(P<0.05),while downregulation of SRSF6 inhibited their expression(P<0.05).Additionally,the activation of the nuclear factor-κB(NF-κB)and mitogen-activated protein kinase(MAPK)signaling pathways was suppressed by SRSF6 knockdown(P<0.05),indicating that SRSF6 is involved in regulating inflammatory responses in macrophages.Myeloid differentiation factor 88(MyD88)is a key adaptor protein in the TLR4 signaling pathway,with its splicing isoforms MyD88-L and MyD88-S exerting pro-inflammatory and anti-inflammatory effects,respectively.Analysis of RNA-binding protein database and RNA immunoprecipitation showed that SRSF6 binds to MyD 88 mRNA.Splicing analysis indicated that downregulation of SRSF6 promoted the expression of the anti-inflammatory MyD88-S mRNA isoform(P<0.01).Moreover,knockdown of MyD88-S could rescue the expression of inflammatory factors suppressed by SRSF6 downregulation.These findings suggest that SRSF6 regulates MyD 88 alternative splicing in macrophages,thereby affecting the activation of inflammatory signaling pathways and the expression of inflammatory fact

关 键 词：丝氨酸/精氨酸富集剪接因子6 巨噬细胞 炎症反应 髓系分化因子88 选择性剪接 

分 类 号：Q257[生物学—细胞生物学]