miR155 Enhances Autophagy by Reducing MBD5 and METTL3 in Human Liver Cancer Stem Cells

Abstract

Background: miR-155 is a widely reported carcinogenic miRNA, with up-regulated expression in a variety of human cancers. Methods: Liver cancer stem cells were isolated from Huh7 cells; gene infection, RT-PCR, Western blotting and tumorigenesis test in vitro and in vivo were performed to analyze the signaling pathway. Results: we demonstrate that miR-155 inhibits the expression of MBD5 and METTL3 , reducing the expression of DNMT1. In particular, miR155 inhibits the interaction between MBD5 and DNMT1, thereby inhibiting the binding capacity among MBD5, DNMT1 and IGFII (H19 ICR, IGFII DMR, IGFII Enhancer) DNA, inhibiting IGFII DNA methylation modification. Importantly, miR-155 promotes the formation of DNA loops in the IGFII DNA region (H19 ICR-IGFII Enhancer, IGFII DMR-IGFII Enhancer). Therefore, miR155 promotes the binding ability of H19-pre miR675 to histone methyltransferases SUV39h2 dependent on the H19 ICR-IGFII enhancer DNA loop and enhances the methylation modification of histone H3 on the ninth lysine . Then, the binding ability of RNA polII to P300 was enhanced by the formation of H3K9me2-RNA polII-P300 complex. Furthermore, miR155 promotes the binding ability of the RNA PolII-P300 complex to the IGFII promoter dependent on the IGFII DMR-IGFII enhancer DNA loop and promotes the expression of IGFII. Further research shows that miR155 promotes the binding of IGFII to stemness factors and the loading of stemness factors into the promoter region of HGF, which promotes the expression of hepatocyte growth factor HGF. In particular, miR155 promotes the expression of albumin gene (ALB) via HGF-c-MET, and then promotes the Sirt1 expression through ALB. Strikingly, miR155 promotes the autophagy dependent on acetylase Sirt1,including that miR155 promotes LC3 activation dependent on acetylase sirt1 and enhances binding capacity of LC3 to TP53INP2 / DOR, ATG4, ATG3 and ATG7, and the binding ability of the interaction between ATG5, ATG12, ATG6L1, ATG9. More meaningfully, miR155 enhances the ability of H-Ras to bind to the autophagosomes Beclin1, vps34, vps5, Uvrage, bif1, Lamp1, Lamp2, Lamp3, Rubicon, Rab24 dependent on autophagy, thereby enhancing the expression of H-Ras. Furthermore, miR155 enhances the expression of pRaf1, pMEK1 / 2, pERK1 / 2, pElK, pJak, Jun, pAKT, pmTOR, P70S6K, 4E-BP1, SGK1, C-myc dependent on H-Ras.Finally, miR-155 promotes the malignant growth of human liver cancer stem cells mediated by H-Ras.Conclusions: these results provide a valuable theoretical basis for therapeutic targets of liver cancer based on miR-155.