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MRNA aging shapes the Cap2 methylome in mammalian mRNA

ABSTRACT: The mRNA cap structure is a major site of dynamic mRNA methylation. mRNA caps exist in either the Cap1 or Cap2 form, depending on the presence of 2’-O-methylation on the first, or both the first and second transcribed nucleotide, respectively. However, the identity of Cap2-containing mRNAs and the function of Cap2 are unclear. Here we describe CLAM-Cap-Seq, a method for transcriptome-wide mapping and quantification of Cap2. We find that unlike other epitranscriptomic modifications, Cap2 can occur on all mRNAs. Cap2 is formed through a slow continuous conversion of mRNAs from Cap1 to Cap2 as mRNAs age in the cytosol. As a result, Cap2 is enriched on long-lived mRNAs. We find that large increases in the abundance of Cap1 leads to activation of RIG-I, especially in conditions where RIG-I expression is increased. The methylation of Cap1 to Cap2 markedly reduces the ability of RNAs to bind and activate RIG-I. We find that the slow Cap2 methylation rate allows Cap2 to accumulate on host mRNAs yet ensures that low Cap2 levels occur on newly expressed viral RNAs. Overall, these results reveal an immunostimulatory role for Cap1, and that Cap2 functions to reduce activation of the innate immune response.

ORGANISM(S): Mus musculus Danio rerio Homo sapiens Caenorhabditis elegans Drosophila melanogaster

PROVIDER: GSE196043 | GEO | 2022/12/12

REPOSITORIES: GEO

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2'-O-methylation of the second transcribed nucleotide in eukaryotic RNA 5’ cap impacts protein production levels and susceptibility to DXO-mediated transcript degradation

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2022-09-08 | PXD028636 | Pride

Part 2 JAWS; 2'-O-methylation of the second transcribed nucleotide in eukaryotic RNA 5’ cap impacts protein production levels and susceptibility to DXO-mediated transcript degradation

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High-resolution mapping reveals a conserved, widespread, dynamic meiotically regulated mRNA methylation program [Mm]

Project description:N6-methyladenosine (m6A) is a common modification of mRNA, with potential roles in fine-tuning the RNA life cycle, but little is known about the pathways regulating this process and its physiological role. Here, we used mass-spectrometry to identify a dense network of proteins physically interacting with METTL3, a core component of the methyltransferase complex, and show that two of them, WTAP and KIAA1429, are required for methylation. Combining high resolution m6A-Seq with knockdown of WTAP allowed us to define accurate maps, at near single-nucleotide resolution, of sites of mRNA methylation across four dynamic programs in human and mouse, including development, differentiation, reprogramming and immune response. Internal WTAP-dependent methylation sites were largely static across the different surveyed conditions and present in the majority of mRNAs. However, methylations were found at much lower levels within highly expressed mRNAs, and methylation is inversely correlated with mRNA stability, consistent with a role in establishing an overall basal, cell-type invariant, distribution of degradation rates. In addition, we identify thousands of WTAP-independent methylation sites at transcription initiation sites, forming part of the mRNA cap structure. We show that the methylations occur at the first transcribed nucleotide, and find that thousands of transcripts are present in different isoforms differing in the methylation state of the first transcribed nucleotide, a previously unappreciated complexity of the transcriptome. Together, our data sheds new light on the proteomic and transcriptional underpinnings of this epitranscriptomic modification in mammals. Examination of m6A methylation across different knockdowns using shRNAs in mouse embryonic fibroblasts, in embyronic and adult brains, and in dendritic cell stimulated with LPS.

2014-07-18 | E-GEOD-55575 | biostudies-arrayexpress

High-resolution mapping reveals a conserved, widespread, dynamic meiotically regulated mRNA methylation program [Hs]

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