Pseudouridinylation of mRNA coding sequences alters translation

Eyler, Daniel E.; Franco, Monika K.; Batool, Zahra; Wu, Monica Z.; Dubuke, Michelle L.; Dobosz-Bartoszek, Malgorzata; Jones, Joshua D.; Polikanov, Yury S.; Roy, Bijoyita; Koutmou, Kristin S.

doi:10.1073/pnas.1821754116

Pseudouridinylation of mRNA coding sequences alters translation

Journal Article · Thu Oct 31 00:00:00 EDT 2019 · Proceedings of the National Academy of Sciences of the United States of America

DOI:https://doi.org/10.1073/pnas.1821754116· OSTI ID:1575030

^[1]; Franco, Monika K. ^[1]; Batool, Zahra ^[2]; Wu, Monica Z. ^[3]; Dubuke, Michelle L. ^[4]; Dobosz-Bartoszek, Malgorzata ^[2]; Jones, Joshua D. ^[1]; ^[2]; Roy, Bijoyita ^[3]; Koutmou, Kristin S. ^[1]

Univ. of Michigan, Ann Arbor, MI (United States)
Univ. of Illinois, Chicago, IL (United States)
New England Biolabs Inc., Ipswich, MA (United States)
Univ. of Massachusetts, Worcester, MA (United States)

Chemical modifications of RNAs have long been established as key modulators of nonprotein-coding RNA structure and function in cells. There is a growing appreciation that messenger RNA (mRNA) sequences responsible for directing protein synthesis can also be posttranscriptionally modified. The enzymatic incorporation of mRNA modifications has many potential outcomes, including changing mRNA stability, protein recruitment, and translation. In this work, we tested how one of the most common modifications present in mRNA coding regions, pseudouridine (Ψ), impacts protein synthesis using a fully reconstituted bacterial translation system and human cells. Our work reveals that replacing a single uridine nucleotide with Ψ in an mRNA codon impedes amino acid addition and EF-Tu GTPase activation. A crystal structure of the Thermus thermophilus 70S ribosome with a tRNA^Phe bound to a ΨUU codon in the A site supports these findings. We also find that the presence of Ψ can promote the low-level synthesis of multiple peptide products from a single mRNA sequence in the reconstituted translation system as well as human cells, and increases the rate of near-cognate Val-tRNA^Val reacting on a ΨUU codon. The vast majority of Ψ moieties in mRNAs are found in coding regions, and our study suggests that one consequence of the ribosome encountering Ψ can be to modestly alter both translation speed and mRNA decoding.

View Accepted Manuscript (DOE)

Research Organization:: Argonne National Laboratory (ANL), Argonne, IL (United States). Advanced Photon Source (APS)

Sponsoring Organization:: National Institute of General Medical Sciences (NIGMS); National Institutes of Health (NIH); New England Biolabs (NEB) Inc.; Rackham Merit Fellowship; USDOE Office of Science (SC); University of Illinois; University of Michigan

Grant/Contract Number:: AC02-06CH11357

OSTI ID:: 1575030

Journal Information:: Proceedings of the National Academy of Sciences of the United States of America, Journal Name: Proceedings of the National Academy of Sciences of the United States of America Journal Issue: 46 Vol. 116; ISSN 0027-8424

Publisher:: National Academy of SciencesCopyright Statement

Country of Publication:: United States

Language:: ENGLISH

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