LncRNA LEF1-AS1 silencing diminishes EZH2 expression to delay hepatocellular carcinoma development by impairing CEBPB-interaction with CDCA7

Cell Cycle. 2020 Apr;19(8):870-883. doi: 10.1080/15384101.2020.1731052. Epub 2020 Mar 16.

Abstract

Hepatocellular carcinoma (HCC) is recognized for its high mortality rate worldwide. Based on intensive studies, long non-coding RNA (lncRNA) expression exerts significant effects on tumor suppression. Herein, we investigated the molecular mechanism of lymphoid enhancer-binding factor-1 antisense RNA 1 (LEF1-AS1) in HCC cells. Microarray-based gene expression analysis was adopted to predict and verify the differentially expressed genes in HCC, which predicted cell division cycle-associated 7 (CDCA7) and LEF1-AS1 to be highly expressed in HCC. The expression of LEF1-AS1, CDCA7, CCAAT/enhancer-binding protein beta (CEBPB) and enhancer of zeste homolog 2 (EZH2) was determined by means of reverse transcription quantitative polymerase chain reaction (RT-qPCR) and western blot analysis. LncMap was used to predict the lncRNA-transcription factor-gene interaction in HCC. ChIP, RIP assay and dual luciferase reporter gene assay were employed to verify the relationship between the transcription factor and gene. Silencing of LEF1-AS1 could downregulate CDCA7 expression through CEBPB. Overexpression of LEF1-AS1, EZH2 and CDCA7 promoted proliferation and invasion in HCC cells. LEF1-AS1 promoted CDCA7 expression to further upregulate EZH2. Tumor formation in nude mice was assessed to verify the experimental results. Silencing of LEF1-AS1 inhibited the growth of tumors in vivo. Collectively, silencing LEF1-AS1 inhibited the proliferation and invasion of HCC cells by down-regulating EZH2 through the CEBPB-CDCA7 signaling pathway, which provides scientific evidence for the treatment of HCC.Abbreviations: HCC: Hepatocellular carcinoma; lncRNA: long non-coding RNA; LEF1-AS1: lymphoid enhancer-binding factor-1 antisense RNA 1; EZH2: enhancer of zeste homolog 2; CDCA7: cell division cycle-associated 7; GEO: Gene Expression Omnibus; NC: negative control; oe: overexpressed; RT-qPCR: reverse transcription quantitative polymerase chain reaction; PBS: phosphate buffered saline; HRP: horseradish peroxidase; OD: optical density; RIP: Radioimmunoprecipitation; ChIP: Chromatin immunoprecipitation; WT: wild type.

Keywords: CDCA7; CEBPB; EZH2; LEF1-AS1; hepatocellular carcinoma; invasion; proliferation; transcription factor.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adult
  • Aged
  • Animals
  • CCAAT-Enhancer-Binding Protein-beta / genetics
  • CCAAT-Enhancer-Binding Protein-beta / metabolism*
  • Carcinogenesis / genetics*
  • Carcinoma, Hepatocellular / genetics
  • Carcinoma, Hepatocellular / metabolism*
  • Cell Proliferation / genetics
  • Enhancer of Zeste Homolog 2 Protein / genetics
  • Enhancer of Zeste Homolog 2 Protein / metabolism*
  • Female
  • Gene Expression Regulation, Neoplastic
  • Gene Silencing*
  • Hep G2 Cells
  • Humans
  • Liver Neoplasms / genetics
  • Liver Neoplasms / metabolism*
  • Lymphoid Enhancer-Binding Factor 1 / genetics*
  • Male
  • Mice
  • Mice, Inbred BALB C
  • Mice, Nude
  • Middle Aged
  • Nuclear Proteins / genetics
  • Nuclear Proteins / metabolism*
  • RNA, Long Noncoding / genetics
  • RNA, Long Noncoding / metabolism*
  • Signal Transduction / genetics*
  • Transfection
  • Tumor Burden / genetics
  • Xenograft Model Antitumor Assays
  • Young Adult

Substances

  • CCAAT-Enhancer-Binding Protein-beta
  • CDCA7 protein, human
  • CEBPB protein, human
  • LEF1 protein, human
  • Lymphoid Enhancer-Binding Factor 1
  • Nuclear Proteins
  • RNA, Long Noncoding
  • EZH2 protein, human
  • Enhancer of Zeste Homolog 2 Protein

Grants and funding

This study was supported by Key Foundation of Jiangxi Provincial Science and Technology Department (No. 20171ACB21064), Youth Science Foundation of Jiangxi Provincial Science and Technology Department (No. 20151BAB205105), Jiangxi Provincial Science and Technology Planning Project (No. 20141BBG70042), and Science Planning Project of Jiangxi Provincial Education Department (No. GJJ14049) .