Decoding mRNA translatability and stability from the 5′ UTR
Open accessPublished version of a preprint
Abstract
Precise control of protein synthesis by engineering sequence elements in 5' untranslated regions (5' UTRs) remains a fundamental challenge. To accelerate our understanding of the cis-regulatory code embedded in 5' UTRs, we devised massively parallel reporter assays from a synthetic messenger RNA library composed of over one million 5' UTR variants. A completely randomized 10-nucleotide sequence preceding an upstream open reading frame (uORF) and downstream GFP drives a broad range of translational outputs and mRNA stability in mammalian cells. While efficient translation protects mRNA from degradation, uORF translation triggers mRNA decay in a UPF1-dependent manner. We also identified translational inhibitory elements with G-quadruplexes as marks for mRNA decay in P-bodies. Unexpectedly, an unstructured A-rich element in 5' UTRs destabilizes mRNAs in the absence of translation, although it enables cap-independent translation. Our results not only identify diverse sequence features of 5' UTRs that control mRNA translatability, but they also reveal ribosome-dependent and ribosome-independent mRNA-surveillance pathways.
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The rate of mRNA degradation also plays a pivotal role in reporter protein accumulation.
“The rate of mRNA degradation also plays a pivotal role in reporter protein accumulation.”
From Identification of novel 5’ UTR variants for enhanced mRNA translation and vaccine immunogenicity (2026) · cites this paper as [36]
The Results report a positive correlation between reporter mRNA half-life and GFP intensity (Fig. 2c), linking mRNA stability with reporter protein accumulation. But the paper chiefly demonstrates that translation affects mRNA stability; it does not establish that the degradation rate itself plays a pivotal causal role in protein accumulation. (LLM, conf=0.78)
Experimental — accuracy not yet measuredActive translation is thought to protect mRNA from degradation.
“On one hand, active translation is thought to protect mRNA from degradation [36].”
From Identification of novel 5’ UTR variants for enhanced mRNA translation and vaccine immunogenicity (2026) · cites this paper as [36]
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Combinatorial optimization of mRNA structure, stability, and translation for RNA-based therapeutics
Leppek, K., Byeon, G. W., Kladwang, W., Wayment-Steele, H. K., Kerr, C. H., Xu, A. F., Kim, D. S., Topkar, V. V., Choe, C., Rothschild, D., et al. (2022) · Nature Communications · doi:10.1038/s41467-022-28776-w · Cited by 399 · see citing works
Combinatorial optimization of mRNA structure, stability, and translation for RNA-based therapeutics
Leppek, K., Byeon, G. W., Kladwang, W., Wayment-Steele, H. K., Kerr, C. H., Xu, A. F., Kim, D. S., Topkar, V. V., Choe, C., Rothschild, D., et al. (2021) · doi:10.1101/2021.03.29.437587 · Cited by 399 · see citing works
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Identification of novel 5’ UTR variants for enhanced mRNA translation and vaccine immunogenicity
Mazunina, E. P., Dzharullaeva, A. S., Kleymenov, D. A., Bykonia, E. N., Kozlova, S. R., Usachev, E. V., Panova, E. A., Konopleva, M. V., Telegina, D. V., Kukushkina-Zvezdova, A. A., et al. (2026) · Scientific Reports · doi:10.1038/s41598-026-59586-5
- Authors
- Jia, L., Mao, Y., Ji, Q., Dersh, D., Yewdell, J. W., & Qian, S.-B.
- Affiliations
- Cornell University (US)
- National Institute of Allergy and Infectious Diseases (US)
- Corresponding author
- Qian, Shu-Bing
- Venue
- Nature Structural & Molecular Biology
- Published
- 2020-07-27
- Volume
- 27
- Issue
- 9
- Pages
- 814-821
- ISSN
- 1545-9985 (ISSN-L)
- 1545-9993
- OpenAlex
- W3045414875
- Cited by
- 248
- References
- 4
- Citations by year
- 2026: 44 · 2025: 59 · 2024: 54 · 2023: 39 · 2022: 21 · 2021: 25 · 2020: 6
- FWCI
- 7.29
- Type
- journal article
- Language
- en
- Open access
- green
- doi.org
- Licence
- Locations
- Nature Structural & Molecular Biology (publishedVersion)
- bioRxiv (Cold Spring Harbor Laboratory) (acceptedVersion, cc-by-nd)
- PubMed (publishedVersion)
- Funders
- Howard Hughes Medical Institute (55108556)
- National Institutes of Health (R01GM1222814)
- National Cancer Institute (R21CA227917)
- National Institute of General Medical Sciences (R01GM122814)
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › RNA and protein synthesis mechanisms
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › RNA Research and Splicing
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › RNA modifications and cancer
- Keywords
- 5' untranslated region, mRNA stability, upstream open reading frames, massively parallel reporter assay, UPF1, mRNA degradation, P-bodies, G-quadruplex, cap-independent translation, cis-regulatory elements
- MeSH terms
- Humans, RNA, Messenger, Protein Biosynthesis, Open Reading Frames, 5' Untranslated Regions, RNA Stability, HEK293 Cells
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