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- W3115570369 abstract "Abstract Upstream open reading frame (uORF) translation disrupts scanning 43S flux on mRNA and modulates main open reading frame (mORF) translation efficiency. Current tools, however, have limited access to ribosome dynamics in both upstream and main ORFs of an mRNA. Here, we develop a new two-color in vitro fluorescence assay, Smart-ORF, that monitors individual uORF and mORF translation events in real-time with single-molecule resolution. We demonstrate the utility of Smart-ORF by applying it to uORF-encoded arginine attenuator peptide (AAP)-mediated translational regulation. The method enabled quantification of uORF and mORF initiation efficiencies, 80S dwell time, polysome formation, and the correlation between uORF and mORF translation dynamics. Smart-ORF revealed that AAP-mediated 80S stalling in the uORF stimulates the uORF initiation efficiency and promotes clustering of slower uORF-translating ribosomes. This technology provides a new tool that can reveal previously uncharacterized dynamics of uORF-containing mRNA translation." @default.
- W3115570369 created "2021-01-05" @default.
- W3115570369 creator A5034804463 @default.
- W3115570369 creator A5044108273 @default.
- W3115570369 creator A5049462275 @default.
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- W3115570369 date "2020-12-16" @default.
- W3115570369 modified "2023-10-12" @default.
- W3115570369 title "Smart-ORF: a single-molecule method for accessing ribosome dynamics in both upstream and main open reading frames" @default.
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- W3115570369 doi "https://doi.org/10.1093/nar/gkaa1185" @default.
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