876 research outputs found

    m6A mRNA demethylase FTO regulates melanoma tumorigenicity and response to anti-PD-1 blockade

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    Melanoma is one of the most deadly and therapy-resistant cancers. Here we show that N6-methyladenosine (m6A) mRNA demethylation by fat mass and obesity-associated protein (FTO) increases melanoma growth and decreases response to anti-PD-1 blockade immunotherapy. FTO level is increased in human melanoma and enhances melanoma tumorigenesis in mice. FTO is induced by metabolic starvation stress through the autophagy and NF-κB pathway. Knockdown of FTO increases m6A methylation in the critical protumorigenic melanoma cell-intrinsic genes including PD-1 (PDCD1), CXCR4, and SOX10, leading to increased RNA decay through the m6A reader YTHDF2. Knockdown of FTO sensitizes melanoma cells to interferon gamma (IFNγ) and sensitizes melanoma to anti-PD-1 treatment in mice, depending on adaptive immunity. Our findings demonstrate a crucial role of FTO as an m6A demethylase in promoting melanoma tumorigenesis and anti-PD-1 resistance, and suggest that the combination of FTO inhibition with anti-PD-1 blockade may reduce the resistance to immunotherapy in melanoma. © 2019, The Author(s)

    METTL3 regulates WTAP protein homeostasis

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    The Wilms tumor 1 (WT1)-associated protein (WTAP) is upregulated in many tumors, including, acute myeloid leukemia (AML), where it plays an oncogenic role by interacting with different proteins involved in RNA processing and cell proliferation. In addition, WTAP is also a regulator of the nuclear complex required for the deposition of N6-methyladenosine (m6A) into mRNAs, containing the METTL3 methyltransferase. However, it is not clear if WTAP may have m6A-independent regulatory functions that might contribute to its oncogenic role. Here, we show that both knockdown and overexpression of METTL3 protein results in WTAP protein upregulation, indicating that METTL3 levels are critical for WTAP protein homeostasis. However, we show that WTAP upregulation is not sufficient to promote cell proliferation in the absence of a functional METTL3. Therein, these data indicate that the reported oncogenic function of WTAP is strictly connected to a functional m6A methylation complex

    The RNA m6A Reader YTHDF2 Is Essential for the Post-transcriptional Regulation of the Maternal Transcriptome and Oocyte Competence.

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    YTHDF2 binds and destabilizes N6-methyladenosine (m6A)-modified mRNA. The extent to which this branch of m6A RNA-regulatory pathway functions in vivo and contributes to mammalian development remains unknown. Here we find that YTHDF2 deficiency is partially permissive in mice and results in female-specific infertility. Using conditional mutagenesis, we demonstrate that YTHDF2 is autonomously required within the germline to produce MII oocytes that are competent to sustain early zygotic development. Oocyte maturation is associated with a wave of maternal RNA degradation, and the resulting relative changes to the MII transcriptome are integral to oocyte quality. The loss of YTHDF2 results in the failure to regulate transcript dosage of a cohort of genes during oocyte maturation, with enrichment observed for the YTHDF2-binding consensus and evidence of m6A in these upregulated genes. In summary, the m6A-reader YTHDF2 is an intrinsic determinant of mammalian oocyte competence and early zygotic development

    PBRM1 Cooperates with YTHDF2 to Control HIF-1α Protein Translation

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    PBRM1, a component of the chromatin remodeller SWI/SNF, is often deleted or mutated in human cancers, most prominently in renal cancers. Core components of the SWI/SNF complex have been shown to be important for the cellular response to hypoxia. Here we investigated how PBRM1 controls HIF-1alpha activity. We find that PBRM1 is required for HIF-1alpha transcriptional activity and protein levels. Mechanistically, PBRM1 is important for HIF-1alpha mRNA translation, as absence of PBRM1 results in reduced activly transalting HIF-1alpha mRNA. Interestingly, we find that PBRM1, but not BRG1, interacts with the m6A reader protein YTHDF2. HIF-1alpha mRNA is m6A modified, bound by PBRM1 and YTHDF2. PBRM1 is necessary for YTHDF2 binding to HIF-1alpha mRNA and reduction of YTHDF2 results in reduced HIF-1alpha protein expression in cells. Our results identify a SWI/SNF independent function for PBRM1, interacting with HIF-1alpha mRNA and the epitranscriptome machinery. Furthermore, our results suggests that the epitranscriptome associated proteins play a role in the control of hypoxia signalling pathway

    RNA Methylation and Ythdf Readers in Posttranscriptional Regulation and Development

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    Development in animals requires precise and coordinated changes in gene expression. This genetic remodeling is achieved through extensive regulatory networks of proteins and RNAs that function together to specify new cell fates and patterns. One developmental event heavily reliant on these regulatory networks is the maternal-to-zygotic transition (MZT), a universal step in metazoan embryogenesis in which a fertilized oocyte is reprogrammed into a pluripotent embryo. The earliest stages of the MZT are governed by maternally inherited gene products, which are required for cellular functions in the initially transcriptionally silent embryo. To shift developmental control to the zygote, these maternal mRNAs are massively degraded through multiple posttranscriptional mechanisms. The RNA modification, N6-methyladenosine (m6A) has been proposed as a master regulator of mRNA decay during developmental transitions, but the direct effects of this pathway on maternal transcript clearance remain unclear. To determine whether m6A facilitates gene expression changes during the MZT, I employed zebrafish embryos as a model system to dissect the contributions of RNA methylation and its reader proteins to maternal transcript fate. Through transcriptome analysis and reporter assays, I found that m6A controls maternal mRNA degradation by promoting deadenylation. To understand how RNA methylation fits into the framework of known decay pathways, I compared transcripts co-targeted by m6A and miR-430, a microRNA that controls mRNA clearance in zebrafish. This revealed that these mechanisms function independently but additively to promote mRNA degradation, reflecting that methylation modulates transcript abundance in concert with known regulators. To disentangle the roles of the Ythdf proteins that mediate the effects of m6A on mRNA, I generated zebrafish genetic mutants of Ythdf1, Ythdf2, and Ythdf3. Through transcriptomic and phenotypic analysis of these mutants, I determined that global maternal mRNA clearance, zygotic genome activation, and development proceed normally in the absence of any one reader. This revealed that individual Ythdf protein have limited effects on the removal of methylated maternal mRNAs during the MZT. To test if this restricted impact of single Ythdf loss stems from functional redundancy between the readers, I produced double mutants of Ythdf2 and Ythdf3. Double Ythdf deletion prevents female gonad development, indicating that these factors exert overlapping activities during oogenesis. Finally, to fully establish functionally redundancy, I created triple Ythdf mutants, which were larval lethal. I observed this same phenotype in zebrafish lacking the methylases that add m6A to mRNA, indicating that RNA methylation is essential for developmental viability. Together, this work provides insight into the contributions of the m6A modification and its Ythdf effectors to maternal mRNA clearance, and establishes how these key regulators coordinate the gene expression changes that underlie embryonic reprogramming

    Genetic Regulation of N6-Methyladenosine-RNA in Mammalian Gametogenesis and Embryonic Development

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    Emerging evidence shows that m(6)A is the most abundant modification in eukaryotic RNA molecules. It has only recently been found that this epigenetic modification plays an important role in many physiological and pathological processes, such as cell fate commitment, immune response, obesity, tumorigenesis, and relevant for the present review, gametogenesis. Notably the RNA metabolism process mediated by m(6)A is controlled and regulated by a series of proteins termed writers, readers and erasers that are highly expressed in germ cells and somatic cells of gonads. Here, we review and discuss the expression and the functional emerging roles of m(6)A in gametogenesis and early embryogenesis of mammals. Besides updated references about such new topics, readers might find in the present work inspiration and clues to elucidate epigenetic molecular mechanisms of reproductive dysfunction and perspectives for future research

    Readers of the m6A epitranscriptomic code

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    International audienceN6-methyl adenosine (m6A) is the most prevalent and evolutionarily conserved, modification of polymerase II transcribed RNAs. By post-transcriptionally controlling patterns of gene expression, m6A deposition is crucial for organism reproduction, development and likely stress responses. m6A mostly mediates its effect by recruiting reader proteins that either directly accommodate the modified residue in a hydrophobic pocket formed by their YTH domain, or otherwise have their affinity positively influenced by the presence of m6A. We firstly describe here the evolutionary history, and review known molecular and physiological roles of eukaryote YTH readers. In the second part, we present non YTH-proteins whose roles as m6A readers largely remain to be explored. The diversity and multiplicity of m6A readers together with the possibility to regulate their expression and function in response to various cues, offers a multitude of possible combinations to rapidly and finely tune gene expression patterns and hence cellular plasticity
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