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N ⁶-methyladenosine of chromosome-associated regulatory RNA regulates chromatin state and transcription.

ABSTRACT: N ⁶-methyladenosine (m⁶A) regulates stability and translation of messenger RNA (mRNA) in various biological processes. In this work, we show that knockout of the m⁶A writer Mettl3 or the nuclear reader Ythdc1 in mouse embryonic stem cells increases chromatin accessibility and activates transcription in an m⁶A-dependent manner. We found that METTL3 deposits m⁶A modifications on chromosome-associated regulatory RNAs (carRNAs), including promoter-associated RNAs, enhancer RNAs, and repeat RNAs. YTHDC1 facilitates the decay of a subset of these m⁶A-modified RNAs, especially elements of the long interspersed element-1 family, through the nuclear exosome targeting-mediated nuclear degradation. Reducing m⁶A methylation by METTL3 depletion or site-specific m⁶A demethylation of selected carRNAs elevates the levels of carRNAs and promotes open chromatin state and downstream transcription. Collectively, our results reveal that m⁶A on carRNAs can globally tune chromatin state and transcription.

SUBMITTER: Liu J

PROVIDER: S-EPMC7213019 | biostudies-literature | 2020 Jan

REPOSITORIES: biostudies-literature

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N 6-methyladenosine of chromosome-associated regulatory RNA regulates chromatin state and transcription.

Liu Jun J Dou Xiaoyang X Chen Chuanyuan C Chen Chuan C Liu Chang C Xu Meng Michelle MM Zhao Siqi S Shen Bin B Gao Yawei Y Han Dali D He Chuan C

Science (New York, N.Y.) 20200116 6477

N 6-methyladenosine (m6A) regulates stability and translation of messenger RNA (mRNA) in various biological processes. In this work, we show that knockout of the m6A writer Mettl3 or the nuclear reader Ythdc1 in mouse embryonic stem cells increases chromatin accessibility and activates transcription in an m6A-dependent manner. We found that METTL3 deposits m6A modifications on chromosome-associated regulatory RNAs (carRN ...[more]

PMID: 31949099

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Project description:N6-methyladenosine (m6A) modification is a major RNA epigenetic regulatory mechanism. The dynamics of m6A levels in viral genomic RNA and their mRNAs have been shown to have either pro- or antiviral functions, and therefore, m6A modifications influence virus-host interactions. Currently, no reports are available on the effect of m6A modifications in the genome of Peste des petits ruminants virus (PPRV). In the present study, we took PPRV as a model for nonsegmented negative-sense single-stranded RNA viruses and elucidate the role of m6A modification on viral replication. We detected m6A-modified sites in the mRNA of the virus and host cells, as well as the PPRV RNA genome. Further, it was found that the level of m6A modification in host cells alters the viral gene expression. Knockdown of the METTL3 and FTO genes (encoding the m6A RNA modification writer and eraser proteins, respectively) results in alterations of the levels of m6A RNA modifications in the host cells. Experiments using these genetically modified clones of host cells infected with PPRV revealed that both higher and lower m6A RNA modification in the host cells negatively affect PPRV replication. We found that m6A-modified viral transcripts had better stability and translation efficiency compared to the unmodified mRNA. Altogether, from these data, we conclude that the m6A modification of RNA regulates PPRV replication. These findings contribute toward a way forward for developing novel antiviral strategies against PPRV by modulating the dynamics of host m6A RNA modification. IMPORTANCE Peste des petits ruminants virus (PPRV) causes a severe disease in sheep and goats. PPRV infection is a major problem, causing significant economic losses to small ruminant farmers in regions of endemicity. N6-methyladenosine (m6A) is an important RNA modification involved in various functions, including virus-host interactions. In the present study, we used stable clones of Vero cells, having knocked down the genes encoding proteins involved in dynamic changes of the levels of m6A modification. We also used small-molecule compounds that interfere with m6A methylation. This resulted in a platform of host cells with various degrees of m6A RNA modification. The host cells with these different microenvironments were useful for studying the effect of m6A RNA modification on the expression of viral genes and viral replication. The results pinpoint the level of m6A modifications that facilitate the maximum replication of PPRV. These findings will be useful in increasing the virus titers in cultured cells needed for the economical development of the vaccine. Furthermore, the findings have guiding significance for the development of novel antiviral strategies for limiting PPRV replication in infected animals.

Project description:Muscle-specific adult stem cells (MuSCs) are required for skeletal muscle regeneration. To ensure efficient skeletal muscle regeneration after injury, MuSCs must undergo state transitions as they are activated from quiescence, give rise to a population of proliferating myoblasts, and continue either to terminal differentiation, to repair or replace damaged myofibers, or self-renewal to repopulate the quiescent population. Changes in MuSC/myoblast state are accompanied by dramatic shifts in their transcriptional profile. Previous reports in other adult stem cell systems have identified alterations in the most abundant internal mRNA modification, N6-methyladenosine (m6A), conferred by its active writer, METTL3, to regulate cell state transitions through alterations in the transcriptional profile of these cells. Our objective was to determine if m6A-modification deposition via METTL3 is a regulator of MuSC/myoblast state transitions in vitro and in vivo. Using liquid chromatography/mass spectrometry we identified that global m6A levels increase during the early stages of skeletal muscle regeneration, in vivo, and decline when C2C12 myoblasts transition from proliferation to differentiation, in vitro. Using m6A-specific RNA-sequencing (MeRIP-seq), a distinct profile of m6A-modification was identified, distinguishing proliferating from differentiating C2C12 myoblasts. RNAi studies show that reducing levels of METTL3, the active m6A methyltransferase, reduced global m6A levels and forced C2C12 myoblasts to prematurely differentiate. Reducing levels of METTL3 in primary mouse MuSCs prior to transplantation enhanced their engraftment capacity upon primary transplantation, however their capacity for serial transplantation was lost. In conclusion, METTL3 regulates m6A levels in MuSCs/myoblasts and controls the transition of MuSCs/myoblasts to different cell states. Furthermore, the first transcriptome wide map of m6A-modifications in proliferating and differentiating C2C12 myoblasts is provided and reveals a number of genes that may regulate MuSC/myoblast state transitions which had not been previously identified.

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N ⁶-methyladenosine of chromosome-associated regulatory RNA regulates chromatin state and transcription.

Publications

<i>N</i> <sup>6</sup>-methyladenosine of chromosome-associated regulatory RNA regulates chromatin state and transcription.

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