Post-transcriptional regulation of BRG1 by FIRΔexon2 in gastric cancer
Post-transcriptional regulation of BRG1 by FIRΔexon2 in gastric cancer
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DOI：
10.1038/s41389-020-0205-4
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发表时间：
2020-02-18
期刊：
影响因子：
6.2
通讯作者：
Matsushita, Kazuyuki
中科院分区：
文献类型：
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作者：
Ailiken, Guzhanuer;Kitamura, Kouichi;Matsushita, Kazuyuki
Brahma相关基因1（BRG 1）是SWItch/蔗糖非发酵（SWI/SNF）染色质重塑复合物的ATP酶亚基，通过三磷酸腺苷依赖的染色体解旋酶DNA结合蛋白7（CHD 7）调控上皮-间质转化（EMT）相关基因，控制多能神经嵴的形成。BRG 1的表达通过癌症中相互作用的RNP参与前mRNA剪接;然而，BRG 1和CHD 7如何与癌症发展相关的详细分子病理学仍然在很大程度上未被揭示。这项研究证明了BRG 1在EMT中的新的转录后调节以及与FIR Delta外显子2的关系，FIR Delta外显子2是缺少外显子2的远上游元件结合蛋白（FUBP）1相互作用阻遏物（FIR）的剪接变体，其不能抑制癌症中的c-myc转录。以前，我们已经报道了FIR完全敲除小鼠（FIR-/-）在E9.5之前是胚胎致死的，这表明FIR对发育至关重要。FIR Delta外显子2通过CHIP序列乙酰化BRG 1启动子上的H3 K27，并抑制BRG 1的转录后表达;在此，BRG 1抑制Snai 1，Snai 1是E-钙粘蛋白的转录抑制因子，其防止癌症侵袭和转移。核糖体蛋白、hnRNP、剪接相关因子、poly（A）结合蛋白、mRNA结合蛋白、tRNA、DEAD盒和WD重复蛋白通过重做穷举质谱分析被鉴定为与FIR和FIR Delta外显子2共免疫沉淀的蛋白。此外，FIR Delta外显子2对FGF 8 mRNA剪接的影响被检查作为CHARGE综合征中显示的受损CHD 7引起的神经发育的指标。预期的是，FIR Delta外显子2的siRNA改变了FGF 8前体mRNA的剪接，表明FIR Delta外显子2、BRG 1和CHD 7之间存在密切的分子相互作用。FIR Delta外显子2 mRNA在人胃癌中升高，但在FIR+/小鼠（K19-Wnt 1/C2mE x FIR+/-）的非侵袭性胃肿瘤中未升高。与野生型小鼠相比，FIR+/-小鼠胃肿瘤中FIR家族（FIR、FIR Delta外显子2和PUF 60）、BRG 1、Snai 1、FBW 7、E-钙粘蛋白、c-Myc、细胞周期蛋白-E和SAP 155的水平增加。FIR家族、Snai 1、细胞周期蛋白-E、BRG 1和c-Myc在Gan小鼠中显示出在较大肿瘤中比在较小肿瘤中更高表达的趋势（K19-Wnt 1/C2mE）。BRG 1和Snai 1在Gan小鼠胃肿瘤中的表达呈正相关。最后，三维晶体结构分析表明，BRG 1是F-box和WD-重复结构域7（FBW 7）的候选底物，FIR Delta外显子2的U2 AF-同源基序（UHM）作为UHM-配体基序与FBW 7降解决定子口袋中的色氨酸-425和WD-样基序相互作用。FIR Delta外显子2共同参与BRG 1的多步转录后调节，通过BRG 1/Snai 1/E-cadherin途径影响EMT，促进肿瘤增殖和胃癌侵袭。
Brahma-related gene 1 (BRG1), an ATPase subunit of the SWItch/sucrose non-fermentable (SWI/SNF) chromatin remodeling complex controls multipotent neural crest formation by regulating epithelial-mesenchymal transition (EMT)-related genes with adenosine triphosphate-dependent chromodomain-helicase DNA-binding protein 7 (CHD7). The expression of BRG1 engages in pre-mRNA splicing through interacting RNPs in cancers; however, the detailed molecular pathology of how BRG1and CHD7 relate to cancer development remains largely unveiled. This study demonstrated novel post-transcriptional regulation of BRG1 in EMT and relationship with FIR Delta exon2, which is a splicing variant of the far-upstream element-binding protein (FUBP) 1-interacting repressor (FIR) lacking exon 2, which fails to repress c-myc transcription in cancers. Previously, we have reported that FIR complete knockout mice (FIR-/-) was embryonic lethal before E9.5, suggesting FIR is crucial for development. FIR Delta exon2 acetylated H3K27 on promoter of BRG1 by CHIP-sequence and suppressed BRG1 expression post-transcriptionally; herein BRG1 suppressed Snai1 that is a transcriptional suppressor of E-cadherin that prevents cancer invasion and metastasis. Ribosomal proteins, hnRNPs, splicing-related factors, poly (A) binding proteins, mRNA-binding proteins, tRNA, DEAD box, and WD-repeat proteins were identified as co-immunoprecipitated proteins with FIR and FIR Delta exon2 by redoing exhaustive mass spectrometry analysis. Furthermore, the effect of FIR Delta exon2 on FGF8 mRNA splicing was examined as an indicator of neural development due to impaired CHD7 revealed in CHARGE syndrome. Expectedly, siRNA of FIR Delta exon2 altered FGF8 pre-mRNA splicing, indicated close molecular interaction among FIR Delta exon2, BRG1 and CHD7. FIR Delta exon2 mRNA was elevated in human gastric cancers but not in non-invasive gastric tumors in FIR+/ mice (K19-Wnt1/C2mE x FIR+/-). The levels of FIR family (FIR, FIR Delta exon2 and PUF60), BRG1, Snai1, FBW7, E-cadherin, c-Myc, cyclin-E, and SAP155 increased in the gastric tumors in FIR+/- mice compared to those expressed in wild-type mice. FIR family, Snai1, cyclin-E, BRG1, and c-Myc showed trends toward higher expression in larger tumors than in smaller tumors in Gan-mice (K19-Wnt1/C2mE). The expressions of BRG1 and Snai1 were positively correlated in the gastric tumors of the Gan-mice. Finally, BRG1 is a candidate substrate of F-box and WD-repeat domain-containing 7 (FBW7) revealed by three-dimensional crystal structure analysis that the U2AF-homology motif (UHM) of FIR Delta exon2 interacted with tryptophan-425 and asparate-399 (WD)-like motif in the degron pocket of FBW7 as a UHM-ligand motif. Together, FIR Delta exon2 engages in multi-step post-transcriptional regulation of BRG1, affecting EMT through the BRG1/Snai1/E-cadherin pathway and promoting tumor proliferation and invasion of gastric cancers.