Identification of Prognostic Factors Related to Super Enhancer-Regulated ceRNA Network in Metastatic Lung Adenocarcinoma.

INTRODUCTION

The regulatory mechanisms of super enhancers (SEs) and ceRNA networks in LUAD progression are not well understood. We aimed to discover the prognostic-related ceRNA network regulated by SEs in metastatic LUAD.

METHODS

RNA-seq data were extracted from The Cancer Genome Atlas (TCGA) database. Differentially expressed (DE) RNAs were identified by edgeR. CeRNA network was predicted and visualized using starBase and Cytoscape. H3K27ac ChIP-seq data were derived from the Gene Expression Omnibus (GEO) database, and used for SE identification. Kaplan-Meier curve and multivariate Cox model were applied for prognostic analysis. Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG) and protein-protein interaction (PPI) network were performed for functional analysis. SEs of AC074117.1 were verified by ChIP-qPCR in A549 and H1299 cells. MTT assay was performed to analyze cell proliferation. Luciferase activity assay was carried out to validate the target targeting relationships of ceRNA network.

RESULTS

A total of 2355 DEmRNA, 483 DElncRNA and 155 DEmiRNA were identified between metastatic LUAD and adjacent normal tissues. CeRNA network consisting of 7 DElncRNAs, 18 DEmiRNAs and 15 DEmRNAs was constructed. Among the seven DElncRNAs in ceRNA network, only AC074117.1 was regulated by SEs. SE-regulated prognostic ceRNA sub-network consisting of FKBP3, E2F2, AC074117.1 and hsa-let-7c-5p was screened and verified. The overlapping co-expressed mRNAs of FKBP3, E2F2, AC074117.1 and hsa-let-7c-5p were mainly related to cell division and Fanconi anemia pathway. Genes in the ceRNA sub-network were correlated with DNA mismatch repair markers. Functional experiments proved that AC074117.1 was highly expressed in LUAD cells. AC074117.1 silencing notably inhibited proliferation of A549 and H1299 cells. Luciferase activity assay confirmed the direct relationship in AC074117.1-hsa-let-7c-5p-FKBP3/E2F2 network.

CONCLUSION

A novel prognostic ceRNA sub-network regulated by SEs was identified in metastatic LUAD. This study provided potential therapeutic targets and prognostic markers for further study of metastatic LUAD.