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METTL3-Mediated N6-Methyladenosine Modification Is Involved in the Dysregulation of NRIP1 Expression in Down Syndrome.

ABSTRACT: Down syndrome (DS) is a common genetic condition in which a person is born with an extra copy of chromosome 21. Intellectual disability is the most common characteristic of DS. N6-methyladenosine (m6A) is a common RNA modification that is implicated in many biological processes. It is highly enriched within the brain and plays an essential role in human brain development. However, the mRNA m6A modification in the fetal brain of DS has not been explored. Here, we report m6A mRNA profiles and mRNA expression profiles of fetal brain cortex tissue from DSs and controls. We observed that the m6A modification in DS brain tissues was reduced genome-wide, which may be due to decreased the m6A methyltransferase like 3 (METTL3) protein expression. The nuclear receptor-interacting protein 1 (NRIP1/RIP140) is coded by a highly conserved chromosome 21 (Hsa21) gene. Overexpression of NRIP1 is associated with mitochondrial dysfunction in DS. The NRIP1 mRNA increased in fetal brain tissues of DS, whereas the m6A modification of the NRIP1 mRNA significantly decreased. METTL3 knockdown reduced the m6A modification of NRIP1 mRNA and increased its expression, and an increase in NRIP1 m6A modification and a decrease in its expression were observed in METTL3-overexpressed cells. The Luciferase reporter assay confirmed that METTL3 regulates NRIP1 expression in an m6A-dependent manner. The decay rate of NRIP1 mRNA was significantly reduced in METTL3-knockdown cells but increased in METTL3-overexpressed cells. We proposed that the m6A modification of NRIP1 mRNA in DS fetal brain tissue is reduced, reducing its transcript degradation rate, resulting in abnormally increased expression of NRIP1, at least partially, in the DS brain. It provides a new mechanism for the molecular pathology of DS and leads to a new insight that may become therapeutically relevant.

SUBMITTER: Shi W

PROVIDER: S-EPMC8047211 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BackgroundAs one of the most frequent chemical modifications in eukaryotic mRNAs, N6-methyladenosine (m6A) modification exerts important effects on mRNA stability, splicing, and translation. Recently, the regulatory role of m6A in tumorigenesis has been increasingly recognized. However, dysregulation of m6A and its functions in tumor epithelial-mesenchymal transition (EMT) and metastasis remain obscure.MethodsqRT-PCR and immunohistochemistry were used to evaluate the expression of methyltransferase-like 3 (METTL3) in gastric cancer (GC). The effects of METTL3 on GC metastasis were investigated through in vitro and in vivo assays. The mechanism of METTL3 action was explored through transcriptome-sequencing, m6A-sequencing, m6A methylated RNA immunoprecipitation quantitative reverse transcription polymerase chain reaction (MeRIP qRT-PCR), confocal immunofluorescent assay, luciferase reporter assay, co-immunoprecipitation, RNA immunoprecipitation and chromatin immunoprecipitation assay.ResultsHere, we show that METTL3, a major RNA N6-adenosine methyltransferase, was upregulated in GC. Clinically, elevated METTL3 level was predictive of poor prognosis. Functionally, we found that METTL3 was required for the EMT process in vitro and for metastasis in vivo. Mechanistically, we unveiled the METTL3-mediated m6A modification profile in GC cells for the first time and identified zinc finger MYM-type containing 1 (ZMYM1) as a bona fide m6A target of METTL3. The m6A modification of ZMYM1 mRNA by METTL3 enhanced its stability relying on the "reader" protein HuR (also known as ELAVL1) dependent pathway. In addition, ZMYM1 bound to and mediated the repression of E-cadherin promoter by recruiting the CtBP/LSD1/CoREST complex, thus facilitating the EMT program and metastasis.ConclusionsCollectively, our findings indicate the critical role of m6A modification in GC and uncover METTL3/ZMYM1/E-cadherin signaling as a potential therapeutic target in anti-metastatic strategy against GC.

Project description:Aortic dissection (AD) develops pathological changes in the separation of the true and false aortic lumen, with high lethality. m6A methylation and oxidative stress have also been shown to be involved in the onset of AD. Through bioinformatics methods, three differentially expressed m6A regulators (YTHDC1, YTHDC2, and RBM15) were excavated from the GSE52093 dataset in the Gene Expression Omnibus (GEO) database, and functional enrichment analysis of the differentially expressed genes (DEGs) regulated by m6A regulators was performed. Then, the genes with oxidative stress-related functions among these genes were found. The protein interaction network of the oxidative stress-related genes and the competing endogenous RNA- (ceRNA-) miRNA-mRNA network were constructed. Among them, DHCR24, P4HB, and PDGFRA, which have m6A differences in AD samples, were selected as key genes. We also performed immune infiltration analysis, as well as cell-gene correlation analysis, on samples from the dataset. The results showed that YTHDC1 was positively correlated with macrophage M1 and negatively correlated with macrophage M2. Finally, we extracted AD and healthy aorta RNA and protein from human tissues that were taken from AD patients and patients who received heart transplants, performed quantitative real-time PCR (qRT-PCR) on YTHDC2 and RBM15, and performed qRT-PCR and western blot (WB) detection on YTHDC1 to verify their differences in AD. The mRNA and protein levels of YTHDC1 were consistent with the results of bioinformatics analysis and were downregulated in AD. Immunofluorescence (IF) was used to colocalize YTHDC1 and endothelial cell marker CD31. After knocking down YTHDC1 in human umbilical vein endothelial cells (HUVECs), reactive oxygen species (ROS) levels had a tendency to increase and the expression of peroxide dismutase SOD2 was decreased. This study provides assistance in discovering the role of m6A regulator YTHDC1 in AD. In particular, m6A modification participates in oxidative stress and jointly affects AD.

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METTL3-Mediated N6-Methyladenosine Modification Is Involved in the Dysregulation of NRIP1 Expression in Down Syndrome.

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