N1-methylpseudouridylation of mRNA causes+1 ribosomal frameshifting

• 成果类型: Article
• 署名作者: Mulroney, Thomas E.; Poyry, Tuija; Yam-Puc, Juan Carlos; Rust, Maria; Harvey, Robert F.; Kalmar, Lajos; Horner, Emily; Booth, Lucy; Ferreira, Alexander P.; Stoneley, Mark; Sawarkar, Ritwick; Mentzer, Alexander J.; Lilley, Kathryn S.; Smales, C. Mark; von der Haar, Tobias; Turtle, Lance; Dunachie, Susanna; Klenerman, Paul; Thaventhiran, James E. D.; Willis, Anne E.
• 署名单位: University of Cambridge; University of Oxford; Wellcome Centre for Human Genetics; University of Cambridge; University of Kent; University College Dublin; University of Liverpool; Oxford University Hospitals NHS Foundation Trust; University of Oxford; University of Oxford; Mahidol Oxford Tropical Medicine Research Unit (MORU); Mahidol University; University of Oxford
• 刊物名称: Nature
• ISSN/ISSBN: 0028-6418
• DOI: 10.1038/s41586-023-06800-3
• 发表日期: 2024-01-04
• 关键词: enhances translation; immunogenicity; pseudouridine; nucleotide; sequence; site

摘要

In vitro-transcribed (IVT) mRNAs are modalities that can combat human disease, exemplified by their use as vaccines for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). IVT mRNAs are transfected into target cells, where they are translated into recombinant protein, and the biological activity or immunogenicity of the encoded protein exerts an intended therapeutic effect(1,2). Modified ribonucleotides are commonly incorporated into therapeutic IVT mRNAs to decrease their innate immunogenicity(3-5), but their effects on mRNA translation fidelity have not been fully explored. Here we demonstrate that incorporation of N-1-methylpseudouridine into mRNA results in +1 ribosomal frameshifting in vitro and that cellular immunity in mice and humans to +1 frameshifted products from BNT162b2 vaccine mRNA translation occurs after vaccination. The +1 ribosome frameshifting observed is probably a consequence of N-1-methylpseudouridine-induced ribosome stalling during IVT mRNA translation, with frameshifting occurring at ribosome slippery sequences. However, we demonstrate that synonymous targeting of such slippery sequences provides an effective strategy to reduce the production of frameshifted products. Overall, these data increase our understanding of how modified ribonucleotides affect the fidelity of mRNA translation, and although there are no adverse outcomes reported from mistranslation of mRNA-based SARS-CoV-2 vaccines in humans, these data highlight potential off-target effects for future mRNA-based therapeutics and demonstrate the requirement for sequence optimization.