Dataset Information

Epigenetic regulation of miR-518a-5p-CCR6 feedback loop promotes both proliferation and invasion in diffuse large B cell lymphoma.

ABSTRACT: To investigate the detailed functions and underlying mechanisms of miR-518a-5p/CCR6 in diffuse large B cell lymphoma (DLBCL) is needed. In this study, CCR6 expression levels were tested both in DLBCL cell lines and specimens. Through bioinformatics analysis and quantitative real-time PCR (qRT-PCR) validation, CCR6's targeted miRNA was obtained. Dual luciferase assay was used to verify their targeted relationship. Futhermore, using qRT-PCR, western blot, CCK8, Transwell assays, flow cytometry, pyrosequencing, chromatin immunoprecipitation, and azacitidine/C646 treatment, the detailed functions and underlying mechanisms of CCR6 and its targeted miRNA in DLBCL were detected. We found that negative correlation existed between CCR6 and miR-518a-5p in DLBCL. Both up-regulated miR-518a-5p and down-regulated CCR6 inhibited cell proliferation and invasion in vitro. Experiment then verified the regulatory relationship between miR-518a-5p and CCR6. JAK2 and STAT6 levels were reduced in DLBCL cells transfected with miR-518a-5p mimic or CCR6 small interfering RNA. Interestingly, we showed for the first time that a hyper-methylated condition existed at the promoter region of miR-518a-5p and azacitidine changed levels of miR-518a-5p in a time- and concentration-dependent manner. Finally, we found an enriched histone H3 on lysine 27 acetylation existed in the promoter of CCR6, whose expression could also be changed via C646 in a time- and concentration-dependent manner. The above results suggest that miR-518a-5p-CCR6 feedback loop plays a critical role in DLBCL development. The overexpression of CCR6 is mainly mediated by epigenetic modification through transcriptional and post-transcriptional activation, which provides new directions for DLBCL treatment.

SUBMITTER: Huang Q

PROVIDER: S-EPMC7889247 | biostudies-literature |

REPOSITORIES: biostudies-literature

ACCESS DATA

Similar Datasets

Project description:BACKGROUND:Angiogenesis plays a critical role in the progression of glioma. Previous studies have indicated that RNA-binding proteins (RBPs) interact with RNAs and participate in the regulation of the malignant behaviors of tumors. As a type of endogenous non-coding RNAs, circular RNAs (circRNAs) are abnormally expressed in various cancers and are involved in diverse tumorigeneses including angiogenesis. METHODS:The expression levels of FUS, circ_002136, miR-138-5p, SOX13, and SPON2 were determined using quantitative real-time PCR (qRT-PCR) and western blot. Transient cell transfection was performed using the Lipofectamine 3000 reagent. The RNA-binding protein immunoprecipitation (RNA-IP) and the RNA pull-down assays were used to detect the interaction between FUS and circ_002136. The dual-luciferase reporter assay system was performed to detect the binding sites of circ_002136 and miR-138-5p, miR-138-5p and SOX13. The chromatin immunoprecipitation (ChIP) assays were used to examine the interactions between transcription factor SOX13 and its target proteins . RESULTS:We demonstrated that down-regulation of FUS or circ_002136 dramatically inhibited the viability, migration and tube formation of U87 glioma-exposed endothelial cells (GECs). MiR-138-5p was down-regulated in GECs and circ_002136 functionally targeted miR-138-5p in an RNA-induced silencing complex (RISC). Inhibition of circ_002136, combined with the restoration of miR-138-5p, robustly reduced the angiogenesis of GECs. As a target gene of miR-138-5p, SOX13 was overexpressed in GECs and was proved to be involved in circ_002136 and miR-138-5p-mediated angiogenesis in gliomas. In addition, we found that SOX13 was directly associated with and activated the SPON2 promoter, thereby up-regulating the expression of SPON2 at the transcriptional level. Knockdown of SPON2 suppressed the angiogenesis in GECs. More important, SOX13 activated the FUS promoter and increased its expression, forming a feedback loop. CONCLUSION:Our data suggests that the feedback loop of FUS/circ_002136/miR-138-5p/SOX13 played a crucial role in the regulation of angiogenesis in glioma. This also provides a potential target and an alternative strategy for combined glioma therapy.

Project description:Epigenetic modifications are important early events during carcinogenesis. In particular, hypermethylation of CpG islands in the promoter region of tumor suppressor genes is a well-known mechanism of gene silencing that contributes to cancer development and progression. Tissue factor pathway inhibitor 2 (TFPI-2) is a tumor suppressor involved in invasiveness inhibition. Although TFPI-2 transcriptional silencing, through promoter hypermethylation, has been widely reported in several human malignancies, it has never been explored in lymphoma. In the present study TFPI-2 methylation and gene expression have been investigated in canine Diffuse Large B-cell lymphomas (cDLBCL). The methylation level of 23 CpGs located within the TFPI-2 promoter was investigated by bisulfite-specific PCR and next generation amplicon deep sequencing (GS Junior 454, Roche) in 22 cDLBCLs and 9 controls. For the same specimens, TFPI-2 gene expression was assessed by means of Real-time RT-PCR. Sequence analysis clearly demonstrated that TFPI2 is frequently hypermethylated in cDLBCL. Hypermethylation of the TFPI-2 promoter was found in 77% of DLBCLs (17 out of 22) and in one normal lymph node. Globally, dogs with DLBCL showed a mean methylation level significantly increased compared to controls (p<0.01) and analysis of hypermethylation by site identified 19 loci out of 23 (82%) with mean methylation levels from 2- to 120-fold higher in cDLBCL. Gene expression analysis confirmed a significant down-regulation of TFPI-2 (p<0.05) in DLBCLs compared with normal lymph nodes, suggesting that TFPI-2 hypermethylation negatively regulates its transcription. In addition, a significant positive correlation (p<0.01) was found between TFPI-2 methylation levels and age providing the first indication of age-associated epigenetic modifications in canine DLBCL. To conclude, our findings demonstrated that epigenetic dysregulation of TFPI-2, leading to its reduced expression, is frequently detected in canine DLBCL. In the next future, the aberrant TFPI-2 promoter hypermethylation may be considered in association with prognosis and therapy.

Dataset Information

Epigenetic regulation of miR-518a-5p-CCR6 feedback loop promotes both proliferation and invasion in diffuse large B cell lymphoma.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure