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| Status |
Public on Jul 10, 2024 |
| Title |
Calibrated ribosome profiling assesses the dynamics of ribosomal flux on transcripts |
| Organisms |
Escherichia coli; Arabidopsis thaliana; Saccharomyces cerevisiae; Drosophila melanogaster; Homo sapiens; Mus musculus |
| Experiment type |
Expression profiling by high throughput sequencing
Other
|
| Summary |
Ribosome profiling, which is based on deep sequencing of ribosome footprints, has served as a powerful tool for elucidating the regulatory mechanism of protein synthesis. However, the current method has substantial issues: contamination by rRNAs and the lack of appropriate methods to measure ribosome numbers in transcripts. Here, we overcomeovercame these hurdles through the development of “Ribo-FilterOut”, which is based on the separation of footprints from ribosome subunits by ultrafiltration, and “Ribo-Calibration”, which relies on external spike-ins of stoichiometrically defined mRNA-ribosome complexes. A combination of these approaches estimates the number of ribosomes on a transcript, the translation initiation rate, and the overall number of translation events before its decay, all in a genome-wide manner. Moreover, our method revealed the allocation of ribosomes under heat shock stress, during aging, and across cell types. Our strategy of modified ribosome profiling measures kinetic and stoichiometric parameters of cellular translation across the transcriptome.
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| |
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| Overall design |
Ribosome profiling and RNA-seq of multiple cell lines to detect the global translation of each conditions.
|
| Web link |
https://www.nature.com/articles/s41467-024-51258-0
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| |
|
| Contributor(s) |
Tomuro K, Mito M, Miyake T, Chow SY, Doi M, Ikeuchi Y, Shichino Y, Iwasaki S, Toh H, Kawamoto N |
| Citation(s) |
39187487 |
| |
| Submission date |
May 26, 2023 |
| Last update date |
Oct 09, 2024 |
| Contact name |
Shintaro Iwasaki |
| E-mail(s) |
shintaro.iwasaki@riken.jp
|
| Organization name |
RIKEN
|
| Department |
CPR
|
| Lab |
RNA Systems Biochemistry Laboratory
|
| Street address |
2-1, Hirosawa
|
| City |
Wako city |
| ZIP/Postal code |
351-0198 |
| Country |
Japan |
| |
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| Platforms (9)
|
| GPL24676 |
Illumina NovaSeq 6000 (Homo sapiens) |
| GPL25244 |
Illumina NovaSeq 6000 (Drosophila melanogaster) |
| GPL25368 |
Illumina NovaSeq 6000 (Escherichia coli) |
| GPL26208 |
Illumina NovaSeq 6000 (Arabidopsis thaliana) |
| GPL27812 |
Illumina NovaSeq 6000 (Saccharomyces cerevisiae) |
| GPL34284 |
Illumina NovaSeq X Plus (Homo sapiens) |
|
| Samples (114)
|
|
| Relations |
| BioProject |
PRJNA976655 |
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