Post-transcriptional regulation of BRG1 by FIRΔexon2 in gastric cancer

Abstract

AbstractBrahma-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 FIRΔexon2, 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. FIRΔexon2 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 FIRΔexon2 by redoing exhaustive mass spectrometry analysis. Furthermore, the effect of FIRΔexon2 on FGF8 mRNA splicing was examined as an indicator of neural development due to impaired CHD7 revealed in CHARGE syndrome. Expectedly, siRNA of FIRΔexon2 altered FGF8 pre-mRNA splicing, indicated close molecular interaction among FIRΔexon2, BRG1 and CHD7. FIRΔexon2 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, FIRΔexon2 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 FIRΔexon2 interacted with tryptophan-425 and asparate-399 (WD)-like motif in the degron pocket of FBW7 as a UHM-ligand motif. Together, FIRΔexon2 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.