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MYC-regulated lncRNA NEAT1 promotes B cell proliferation and lymphomagenesis via the miR-34b-5p-GLI1 pathway in diffuse large B-cell lymphoma.

ABSTRACT: Background:LncRNA NEAT1 has been identified as a tumour driver in many human cancers. However, the underlying mechanism of lncRNA NEAT1 in diffuse large B-cell lymphoma (DLBCL) progression is unclear. Methods:The expression levels of NEAT1, GLI1 and miR-34b-5p were detected by RT-qPCR and Western blotting in DLBCL tissues and cell lines. MTT and colony formation assays were performed to examine cell proliferation, while annexin-V staining and TUNEL assays were performed to measure cell apoptosis. The effect of NEAT1, GLI1 and miR-34b-5p on cell cycle-associated proteins was evaluated by Western blotting. Dual-luciferase reporter and RNA immunoprecipitation (RIP) assays were employed to investigate the interaction between NEAT1 and miR-34b-5p or GLI1 and miR-34b-5p. Moreover, chromatin immunoprecipitation (ChIP) was performed to demonstrate the interaction between MYC and NEAT1. Results:NEAT1 and GLI1 were upregulated while miR-34b-5p was downregulated in DLBCL tissues and cell lines compared to normal controls. Knockdown of NEAT1 or overexpression of miR-34b-5p inhibited cell proliferation but promoted cell apoptosis. Overexpression of NEAT1 reversed GLI1-knockdown induced attenuation of cell proliferation. In other words, NEAT1 acted as a competing endogenous RNA (ceRNA), regulating the miR-34b-5p-GLI1 axis, further affecting the proliferation of DLBCL. Moreover, MYC modulated NEAT1 transcription by directly binding to the NEAT1 promoter. Conclusion:We revealed that MYC-regulated NEAT1 promoted DLBCL proliferation via the miR-34b-5p-GLI1 pathway, which could provide a novel therapeutic target for DLBCL.

SUBMITTER: Qian CS

PROVIDER: S-EPMC7081629 | biostudies-literature | 2020

REPOSITORIES: biostudies-literature

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MYC-regulated lncRNA NEAT1 promotes B cell proliferation and lymphomagenesis via the miR-34b-5p-GLI1 pathway in diffuse large B-cell lymphoma.

Qian Chong-Sheng CS Li Ling-Jie LJ Huang Hai-Wen HW Yang Hai-Fei HF Wu De-Pei DP

Cancer cell international 20200319

<h4>Background</h4>LncRNA NEAT1 has been identified as a tumour driver in many human cancers. However, the underlying mechanism of lncRNA NEAT1 in diffuse large B-cell lymphoma (DLBCL) progression is unclear.<h4>Methods</h4>The expression levels of NEAT1, GLI1 and miR-34b-5p were detected by RT-qPCR and Western blotting in DLBCL tissues and cell lines. MTT and colony formation assays were performed to examine cell proliferation, while annexin-V staining and TUNEL assays were performed to measure ...[more]

PMID: 32206038

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Project description:Long-chain non-coding RNAs (LncRNAs) are expressed in diffuse large B-cell lymphoma (DLBCL) tissues and have played a regulatory role in DLBCL with a cancer-promoting effect. In this study, the role of LncRNA SNHG8 in the regulation of DLBCL cells is investigated, and its underlying mechanism is explored. The database of the Gene Expression Profiling Interactive Analysis (GEPIA) was searched, and the expression of SNHG8 in DLBCL and normal tissues was examined. The expression of SNHG8 was evaluated in several DLBCL cell lines and a normal lymphocyte cell line. It was found that SNHG8 was overexpressed in DLBCL tissues and cells in comparison with their normal counterparts. The short hairpin RNA (shRNA) plasmids of SNHG8 were transfected into DLBCL cells to knockdown the expression of SNHG8, followed by assays of proliferation, colony formation, apoptosis, and related protein expression. The results showed that the knockdown of SNHG8 significantly inhibited DLBCL cell proliferation and colony formation while promoting cell apoptosis. Moreover, the knockdown of SNHG8 reduced the expression of Ki-67, proliferating cell nuclear antigen (PCNA), and Bcl-2 and enhanced the expression of Bax and cleaved caspase 3/9. MiR-335-5p was predicted to be a potential target of SNHG8 by using the bioinformatics analysis, and the interaction between the two was validated by using the dual luciferase assay. In addition, the knockdown of SNHG8 increased the level of miR-335-5p, whereas miR-335-5p mimic decreased the expression of SNHG8. Finally, U2932 cells were co-transfected with or without sh-SNHG8 and miR-335-5p inhibitors, whose proliferation, colony formation, and apoptosis were determined subsequently. It was demonstrated that the presence of an miR-335-5p inhibitor partially canceled the inhibitory effects of the knockdown of SNHG8 on DLBCL cell proliferation and colony formation and the stimulating effects of the knockdown of SNHG8 on cell apoptosis. Taken together, our study suggests that lncRNA SNHG8 exerts a cancer-promoting effect on DLBCL via targeting miR-335-5p.

Project description:BackgroundMYC is a heterogeneously expressed transcription factor that plays a multifunctional role in many biological processes such as cell proliferation and differentiation. It is also associated with many types of cancer including the malignant lymphomas. There are two types of aggressive B-cell lymphoma, namely Burkitt lymphoma (BL) and a subgroup of diffuse large cell lymphoma (DLBCL), which both carry MYC translocations and overexpress MYC but both differ significantly in their clinical outcome. In DLBCL, MYC translocations are associated with an aggressive behavior and poor outcome, whereas MYC-positive BL show a superior outcome.MethodsTo shed light on this phenomenon, we investigated the different modes of actions of MYC in aggressive B-cell lymphoma cell lines subdivided into three groups: (i) MYC-positive BL, (ii) DLBCL with MYC translocation (DLBCLpos) and (iii) DLBCL without MYC translocation (DLBCLneg) for control. In order to identify genome-wide MYC-DNA binding sites a chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-Seq) was performed. In addition, ChIP-Seq for H3K4me3 was used for determination of genomic regions accessible for transcriptional activity. These data were supplemented with gene expression data derived from RNA-Seq.ResultsBioinformatics integration of all data sets revealed different MYC-binding patterns and transcriptional profiles in MYC-positive BL and DLBCL cell lines indicating different functional roles of MYC for gene regulation in aggressive B-cell lymphomas. Based on this multi-omics analysis we identified ADGRE5 (alias CD97) - a member of the EGF-TM7 subfamily of adhesion G protein-coupled receptors - as a MYC target gene, which is specifically expressed in BL but not in DLBCL regardless of MYC translocation.ConclusionOur study describes a diverse genome-wide MYC-DNA binding pattern in BL and DLBCL cell lines with and without MYC translocations. Furthermore, we identified ADREG5 as a MYC target gene able to discriminate between BL and DLBCL irrespectively of the presence of MYC breaks in DLBCL. Since ADGRE5 plays an important role in tumor cell formation, metastasis and invasion, it might also be instrumental to better understand the different pathobiology of BL and DLBCL and help to explain discrepant clinical characteristics of BL and DLBCL.

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Dataset Information

MYC-regulated lncRNA NEAT1 promotes B cell proliferation and lymphomagenesis via the miR-34b-5p-GLI1 pathway in diffuse large B-cell lymphoma.

Publications

MYC-regulated lncRNA NEAT1 promotes B cell proliferation and lymphomagenesis via the miR-34b-5p-GLI1 pathway in diffuse large B-cell lymphoma.

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