Integrated Analysis of lncRNA-Associated ceRNA Network Identifies Two lncRNA Signatures as a Prognostic Biomarker in Gastric Cancer.

BACKGROUND: Gastric cancer (GC) is a malignant tumour that originates in the gastric mucosal epithelium and is associated with high mortality rates worldwide. Long noncoding RNAs (lncRNAs) have been identified to play an important role in the development of various tumours, including GC. Yet, lncRNA biomarkers in a competing endogenous RNA network (ceRNA network) that are used to predict survival prognosis remain lacking. The aim of this study was to construct a ceRNA network and identify the lncRNA signature as prognostic factors for survival prediction. METHODS: The lncRNAs with overall survival significance were used to construct the ceRNA network. Function enrichment, protein-protein interaction, and cluster analysis were performed for dysregulated mRNAs. Multivariate Cox proportional hazards regression was performed to screen the potential prognostic lncRNAs. RT-qPCR was used to measure the relative expression levels of lncRNAs in cell lines. CCK8 assay was used to assess the proliferation of GC cells transfected with sh-lncRNAs. RESULTS: Differentially expressed genes were identified including 585 lncRNAs, 144 miRNAs, and 2794 mRNAs. The ceRNA network was constructed using 35 DElncRNAs associated with overall survival of GC patients. Functional analysis revealed that these dysregulated mRNAs were enriched in cancer-related pathways, including TGF-beta, Rap 1, calcium, and the cGMP-PKG signalling pathway. A multivariate Cox regression analysis and cumulative risk score suggested that two of those lncRNAs (LINC01644 and LINC01697) had significant prognostic value. Furthermore, the results indicate that LINC01644 and LINC01697 were upregulated in GC cells. Knockdown of LINC01644 or LINC01697 suppressed the proliferation of GC cells. CONCLUSIONS: The authors identified 2-lncRNA signature in ceRNA regulatory network as prognostic biomarkers for the prediction of GC patient survival and revealed that silencing LINC01644 or LINC01697 inhibited the proliferation of GC cells.

Involvement of Sp1/Sp3 in the activation of the GATA-1 erythroid promoter in K562 cells.

GATA-1 is a hematopoietic transcription factor that is essential for the terminal maturation of proerythroblasts, megakaryocytic cells and mast cells. The erythroid-specific promoter of the human GATA-1 gene directs the high expression of a reporter gene in K562 cells. Multiple putative transcription factor binding sites were identified in the promoter from the -860 to the -1 base pair (bp). For a better understanding of the transcriptional control of human GATA-1 gene expression, we tested the transcriptional activity of a series of deletions from the 5' end of the 860-bp promoter. A region between -221 and -128 bp retains most of the transcriptional activity of the full-length promoter. Deletion of the CGCCC box at -195 bp reduced reporter gene activity to 60.4%. Further deletion of the CACCC box at -173 bp nearly abolished reporter gene expression, indicating that the CACCC box is more critical. In vitro experiments of electrophoretic mobility shifts and in vivo studies using chromatin immuno-precipitation (ChIP) assays show that the Sp1/Sp3 proteins bind the CACCC site in the nuclei of K562 cells. Coincidently, hyperacetylation of histones in the GATA-1 erythroid promoter was also shown by ChIP assay. Co-transfection of Sp1 expression plasmids and plasmids with a wild-type promoter showed enhanced reporter gene activity in a dose-dependent manner. The combined data demonstrate that Sp1/Sp3, but not EKLF, is involved in the activation of the GATA-1 erythroid promoter, and that histones H3 and H4 are highly acetylated in this promoter region for an actively transcribed GATA-1 gene in K562 cells in which EKLF is barely detectable.