title: Incorporation of pseudouridine into mRNA enhances translation by diminishing PKR activation.
abstract: Previous studies have shown that the translation level of in vitro transcribed messenger RNA (mRNA) is enhanced when its uridines are replaced with pseudouridines; however, the reason for this enhancement has not been identified. Here, we demonstrate that in vitro transcripts containing uridine activate RNA-dependent protein kinase (PKR), which then phosphorylates translation initiation factor 2-alpha (eIF-2α), and inhibits translation. In contrast, in vitro transcribed mRNAs containing pseudouridine activate PKR to a lesser degree, and translation of pseudouridine-containing mRNAs is not repressed. RNA pull-down assays demonstrate that mRNA containing uridine is bound by PKR more efficiently than mRNA with pseudouridine. Finally, the role of PKR is validated by showing that pseudouridine- and uridine-containing RNAs were translated equally in PKR knockout cells. These results indicate that the enhanced translation of mRNAs containing pseudouridine, compared to those containing uridine, is mediated by decreased activation of PKR.
authors.affiliation: Department of Medicine, Department of Neurosurgery, University of Pennsylvania, Philadelphia, PA, USA.
authors.forename: Bart R
authors.initials: BR
authors.lastname: Anderson
authors.forename: Hiromi
authors.initials: H
authors.lastname: Muramatsu
authors.forename: Subba R
authors.initials: SR
authors.lastname: Nallagatla
authors.forename: Philip C
authors.initials: PC
authors.lastname: Bevilacqua
authors.forename: Lauren H
authors.initials: LH
authors.lastname: Sansing
authors.forename: Drew
authors.initials: D
authors.lastname: Weissman
authors.forename: Katalin
authors.initials: K
authors.lastname: Karikó
mesh_terms: D000818:Animals; D002460:Cell Line; D002478:Cells, Cultured; D006801:Humans; D051379:Mice; D014176:Protein Biosynthesis; D011560:Pseudouridine / Q000378:metabolism*; D012333:RNA, Messenger / Q000737:chemistry / Q000378:metabolism*; D019892:eIF-2 Kinase / Q000037:antagonists & inhibitors / Q000378:metabolism*
chemical_list: D012333:RNA, Messenger; D011560:Pseudouridine; D019892:eIF-2 Kinase
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