Showing papers on "MRNA modification published in 2013"
TL;DR: The discovery of ALKBH5 as another mammalian demethylase that oxidatively reverses m(6)A in mRNA in vitro and in vivo strongly suggests that the reversible m( 6)A modification has fundamental and broad functions in mammalian cells.
2,274 citations
TL;DR: This article generated genomic maps of m(6)-methyladenosine (m(6)A) sites in meiotic yeast transcripts at nearly single-nucleotide resolution, identifying 1,308 putatively methylated sites within 1,183 transcripts.
529 citations
TL;DR: A polymerase with reverse transcriptase activity that is selective by up to 18-fold for incorporation of thymidine opposite unmodified A over m(6)A is identified and it is demonstrated that the long-undetermined position of m( 6)A in mammalian 28S rRNA is nucleotide 4190.
Abstract: N(6)-methyladenosine (m(6)A) is the most abundant mRNA modification and has important links to human health While recent studies have successfully identified thousands of mammalian RNA transcripts containing the modification, it is extremely difficult to identify the exact location of any specific m(6)A Here we have identified a polymerase with reverse transcriptase activity (from Thermus thermophilus) that is selective by up to 18-fold for incorporation of thymidine opposite unmodified A over m(6)A We show that the enzyme can be used to locate and quantify m(6)A in synthetic RNAs by analysis of pausing bands, and have used the enzyme in tandem with a nonselective polymerase to locate the presence and position of m(6)A in high-abundance cellular RNAs By this approach we demonstrate that the long-undetermined position of m(6)A in mammalian 28S rRNA is nucleotide 4190
82 citations
TL;DR: These variants showed complete compatibility with long‐term expression in mammalian cells, suggesting that they may be usefully applied in functional genomics and genetically modified animal models.
Abstract: ARCHAEA-ExPRESs is an mRNA modification technology that makes use of components derived from the Archaeon Methanocaldococcus jannaschii, namely the tRNA splicing endonuclease (MJ-EndA) and its natural substrate, the bulge-helix-bulge (BHB) structure (1). These components can perform both cis- and trans-splicing in cellular and animal models and may provide a convenient way to modulate gene expression using components independent of cellular regulatory networks. To use MJ-EndA in stable expression mammalian systems, we developed variants characterized by high efficiency and sustainable in vivo activity. The MJ-EndA variants were created by the introduction of proper localization signals followed by mutagenesis and direct selection in mammalian cells. Of note, enzyme selection used an in vivo selection method based on puromycin resistance conferred to cells by BHB-mediated intron splicing from an out-of-frame puromycin N-acetyl transferase (PAC) gene. This approach yielded several endonuclease variants, the...
1 citations