Arginine methylation of METTL14 promotes RNA N6-methyladenosine modification
Ontology highlight
ABSTRACT:
ORGANISM(S): Mus musculus
PROVIDER: GSE164047 | GEO | 2021/05/14
REPOSITORIES: GEO
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Project description:SETD2 is the specific methyltransferase of H3K36me3, while METTL3, METTL14 and WTAP are the components of m6A methyltransferase complex. To understand the global effect of H3K36me3 on m6A modification, we compared the m6A profiling in SETD2 and METTL3, METTL14 or WTAP knockdown HepG2 cells, and found depletion of H3K36me3 by SETD2 silencing globally reduced m6A in human transcriptome. What’s more, most of the SETD2-dependent hypomethylation sites also responded to knockdown of METTL3, METTL14, or WTAP.
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Project description:Chemical modification of RNAs is important for post-transcriptional gene regulation. The METTL3-METTL14 complex generates most N6-methyladenosine (m6A) modifications in mRNAs, and dysregulated methyltransferase expression has been linked to numerous cancers. Here we show that m6A modification location, rather than the overall modification level, can impact oncogenesis. A gain-of-function missense mutation found in cancer patients, METTL14R298P, promotes malignant cell growth in culture and in transgenic mice. The mutant methyltransferase preferentially modifies noncanonical sites and transforms gene expression without increasing global m6A levels in mRNAs. The altered substrate specificity is intrinsic to METTL3-METTL14, helping us to propose a structural model for how the METTL3-METTL14 complex detects RNA sequences. Together, our work highlights that m6A location is important for function and that noncanonical methylation sites may impact aberrant gene expression and oncogenesis.
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Project description:N6-methyladenosine (m6A) methylation of mRNA by the methyltransferase complex (MTC), with core components including METTL3-METTL14 heterodimers and Wilms’ tumor 1-associated protein (WTAP), contributes to breast tumorigenesis, but the mechanism of MTC assembly remains elusive. Here, we identify a novel cleaved form METTL3a (residues 239-580 of METTL3), that is highly expressed in breast cancer. Furthermore, we find that both METTL3a and full-length METTL3 are required for MTC assembly, RNA m6A deposition, as well as cancer cell proliferation. Mechanistically, we find that METTL3a is required for METTL3-METTL3 interaction, which is a prerequisite step for recruitment of WTAP in MTC assembly. Analysis of m6A sequencing data shows that depletion of METTL3a globally disrupts m6A methylation, and METTL3a mediates mTOR activation via m6A-mediated suppression of TMEM127 expression. Consequently, we find that METTL3 cleavage is mediated by proteasome in an mTOR-dependent manner, revealing positive regulatory feedback between METTL3a and mTOR signaling. Our findings reveal METTL3a as an important component for MTC assembly, and suggest the METTL3a-mTOR axis as a potential therapeutic target for breast cancer.
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Project description:METTL3 and METTL14 are considered to faithfully form the m6A writing complex in a 1:1 ratio, regulating the fate of mRNA by adding m6A modifications. However, recent studies have shown inconsistent expression and prognostic value of METTL3 and METTL14 in some tumors, suggesting that they may not be faithful in tumors. Pan-cancer analysis based on TCGA data reveals significant differences in expression, function, tumor burden correlation, and immune correlation between METTL3 and METTL14, especially in esophageal squamous cell carcinoma (ESCC). Knockdown of METTL3 significantly inhibits the cell proliferation in vitro and in vivo in ESCC EC109 cells, while the impact of METTL14 knockdown on proliferation is limited, and it cannot abolish the expression of METTL3 protein. mRNA-seq results indicate that METTL3 independently regulates the expression of 1615 genes, while only 776 genes are co-regulated by METTL3 and METTL14. Furthermore, through immunofluorescence co-localization, it is observed that METTL3 and METTL14 have certain inconsistencies in cellular localization. HPLC-MS results show that METTL3 independently binds to the Nop56p-associated pre-rRNA complex and mRNA splicing complex, separate from METTL14. Through bioinformatics and various omics studies, we have preliminarily discovered that METTL3 independently regulating tumor cell proliferation, and the participation in mRNA splicing may be a critical molecular mechanism. Our study provides an experimental basis and theoretical foundation for further understanding of the m6A writing complex and tumor therapy targeting METTL3.
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Project description:N6-methyladenosine (m6A) is the most abundant internal modification in the messenger RNA (mRNA) of all higher eukaryotes. This modification has been shown to be reversible in mammals; it is installed by a methyltransferase heterodimer complex of METTL3 and METTL14 bound with WTAP, and reversed by iron(II)- and α-ketoglutarate-dependent demethylases FTO and ALKBH5. This modification exhibits significant functional roles in various biological processes. The m6A modification as a RNA mark is recognized by reader proteins, such as YTH domain family proteins and HNRNPA2B1; m6A can also act as a structure switch to affect RNA-protein interactions for biological regulation. In Arabidopsis thaliana, the methyltransferase subunit MTA (the plant orthologue of human METTL3, encoded by At4g10760) was well characterized and FIP37 (the plant orthologue of human WTAP) was first identified as the interacting partner of MTA. Here we report the discovery and characterization of reversible m6A methylation mediated by AtALKBH10B (encoded by At4g02940) in A. thaliana, and noticeable roles of this RNA demethylase in affecting plant development and floral transition. Our findings reveal potential broad functions of reversible mRNA methylation in plants. m6A peaks were identified from wild type Columbia-0 and atalkbh10b-1 mutant in two biological replicates
2016-03-24 | E-GEOD-79523 | biostudies-arrayexpress
Project description:METTL3 and METTL14 are two components that form the core heterodimer of the main RNA m6A methyltransferase complex (MTC, also known as m6A writer) that installs m6A. Surprisingly, depletion of METTL3 or METTL14 displayed distinct effects on mouse embryonic stem cell (mESC) self-renewal. While comparable global hypo-methylation in RNA m6A was observed in Mettl3 or Mettl14 knockout mESCs, respectively. Mettl14 knockout led to a globally decreased nascent RNA synthesis, whereas Mettl3 depletion resulted in transcription upregulation, suggesting that METTL14 might possess an m6A-indenepent role in gene regulation. We found that METTL14 colocalizes with the repressive H3K27me3 modification and PRC2 complex. Mechanically, METTL14, but not METTL3, recognizes H3K27me3 and recruits KDM6B to induce H3K27me3 demethylation independent of METTL3. Depletion of METTL14 thus led to a global increase in H3K27me3 level along with a global gene suppression . The regulation of H3K27me3 by METTL14 is essential to the transition of mESCs from self-renewal to differentiation. This work reveals a regulation mechanism on heterochromatin by METTL14 in a manner distinct from METTL3 and independently of m6A, and critically impacts transcriptional regulation, stemness maintenance and differentiation of mESCs.
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Project description:METTL3 and METTL14 are two components that form the core heterodimer of the main RNA m6A methyltransferase complex (MTC, also known as m6A writer) that installs m6A. Surprisingly, depletion of METTL3 or METTL14 displayed distinct effects on mouse embryonic stem cell (mESC) self-renewal. While comparable global hypo-methylation in RNA m6A was observed in Mettl3 or Mettl14 knockout mESCs, respectively. Mettl14 knockout led to a globally decreased nascent RNA synthesis, whereas Mettl3 depletion resulted in transcription upregulation, suggesting that METTL14 might possess an m6A-indenepent role in gene regulation. We found that METTL14 colocalizes with the repressive H3K27me3 modification and PRC2 complex. Mechanically, METTL14, but not METTL3, recognizes H3K27me3 and recruits KDM6B to induce H3K27me3 demethylation independent of METTL3. Depletion of METTL14 thus led to a global increase in H3K27me3 level along with a global gene suppression . The regulation of H3K27me3 by METTL14 is essential to the transition of mESCs from self-renewal to differentiation. This work reveals a regulation mechanism on heterochromatin by METTL14 in a manner distinct from METTL3 and independently of m6A, and critically impacts transcriptional regulation, stemness maintenance and differentiation of mESCs.
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Project description:METTL3 and METTL14 are two components that form the core heterodimer of the main RNA m6A methyltransferase complex (MTC, also known as m6A writer) that installs m6A. Surprisingly, depletion of METTL3 or METTL14 displayed distinct effects on mouse embryonic stem cell (mESC) self-renewal. While comparable global hypo-methylation in RNA m6A was observed in Mettl3 or Mettl14 knockout mESCs, respectively. Mettl14 knockout led to a globally decreased nascent RNA synthesis, whereas Mettl3 depletion resulted in transcription upregulation, suggesting that METTL14 might possess an m6A-indenepent role in gene regulation. We found that METTL14 colocalizes with the repressive H3K27me3 modification and PRC2 complex. Mechanically, METTL14, but not METTL3, recognizes H3K27me3 and recruits KDM6B to induce H3K27me3 demethylation independent of METTL3. Depletion of METTL14 thus led to a global increase in H3K27me3 level along with a global gene suppression . The regulation of H3K27me3 by METTL14 is essential to the transition of mESCs from self-renewal to differentiation. This work reveals a regulation mechanism on heterochromatin by METTL14 in a manner distinct from METTL3 and independently of m6A, and critically impacts transcriptional regulation, stemness maintenance and differentiation of mESCs.
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Project description:METTL3 and METTL14 are two components that form the core heterodimer of the main RNA m6A methyltransferase complex (MTC, also known as m6A writer) that installs m6A. Surprisingly, depletion of METTL3 or METTL14 displayed distinct effects on mouse embryonic stem cell (mESC) self-renewal. While comparable global hypo-methylation in RNA m6A was observed in Mettl3 or Mettl14 knockout mESCs, respectively. Mettl14 knockout led to a globally decreased nascent RNA synthesis, whereas Mettl3 depletion resulted in transcription upregulation, suggesting that METTL14 might possess an m6A-indenepent role in gene regulation. We found that METTL14 colocalizes with the repressive H3K27me3 modification and PRC2 complex. Mechanically, METTL14, but not METTL3, recognizes H3K27me3 and recruits KDM6B to induce H3K27me3 demethylation independent of METTL3. Depletion of METTL14 thus led to a global increase in H3K27me3 level along with a global gene suppression . The regulation of H3K27me3 by METTL14 is essential to the transition of mESCs from self-renewal to differentiation. This work reveals a regulation mechanism on heterochromatin by METTL14 in a manner distinct from METTL3 and independently of m6A, and critically impacts transcriptional regulation, stemness maintenance and differentiation of mESCs.
2024-08-16 | GSE183396 | GEO