Cellular variability of nonsense-mediated mRNA decay

• Published in: Nature communications 2021-12, Vol.12 (1), p.7203-7203, Article 7203
• Main Authors: Sato, Hanae, Singer, Robert H.
• Format: Article
• Language: English
• Subjects: 13/31; 14; 14/32; 631/1647/2017/1958; 631/337/1645/1769; beta-Globins - genetics; Codon, Nonsense; Codons; Decay; Degradation; Efficiency; Flow Cytometry; Fluorescence; Genes, Reporter - genetics; HEK293 Cells; Heterogeneity; Humanities and Social Sciences; Humans; mRNA turnover; multidisciplinary; Nonsense Mediated mRNA Decay - genetics; Nonsense-mediated mRNA decay; Phosphorylation; Protein Serine-Threonine Kinases; RNA Helicases; RNA, Messenger - metabolism; Science; Science (multidisciplinary); Surveillance; Trans-Activators; Translation termination; Variability
• ISSN: 2041-1723; 2041-1723
• DOI: 10.1038/s41467-021-27423-0

Nonsense-mediated mRNA decay (NMD) is an mRNA degradation pathway that eliminates transcripts containing premature termination codons (PTCs). Half-lives of the mRNAs containing PTCs demonstrate that a small percent escape surveillance and do not degrade. It is not known whether this escape represents variable mRNA degradation within cells or, alternatively cells within the population are resistant. Here we demonstrate a single-cell approach with a bi-directional reporter, which expresses two β-globin genes with or without a PTC in the same cell, to characterize the efficiency of NMD in individual cells. We found a broad range of NMD efficiency in the population; some cells degraded essentially all of the mRNAs, while others escaped NMD almost completely. Characterization of NMD efficiency together with NMD regulators in single cells showed cell-to-cell variability of NMD reflects the differential level of surveillance factors, SMG1 and phosphorylated UPF1. A single-cell fluorescent reporter system that enabled detection of NMD using flow cytometry revealed that this escape occurred either by translational readthrough at the PTC or by a failure of mRNA degradation after successful translation termination at the PTC. Here the author developed a single-cell reporter system to identify cell-to-cell variability of nonsense-mediated mRNA decay (NMD). This approach provides a sensitive tool to investigate cellular heterogeneity of NMD in various physiological conditions.