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| Status |
Public on Nov 14, 2014 |
| Title |
Snapshots of pre-rRNA structural flexibility reveal eukaryotic 40S assembly dynamics at nucleotide resolution |
| Organism |
Saccharomyces cerevisiae |
| Experiment type |
Expression profiling by high throughput sequencing
Other
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| Summary |
Ribosome assembly in eukaryotes involves the activity of hundreds of assembly factors that direct the hierarchical assembly of ribosomal proteins and numerous ribosomal RNA folding steps. However, detailed insights into the function of assembly factors and ribosomal RNA folding events are lacking. To address this, we have developed ChemModSeq, a method that combines structure probing, high throughput sequencing and statistical modeling, to quantitatively measure RNA structural rearrangements during the assembly of macromolecular complexes. By applying ChemModSeq to purified 40S assembly intermediates we obtained nucleotide-resolution maps of ribosomal RNA flexibility revealing structurally distinct assembly intermediates and mechanistic insights into assembly dynamics not readily observed in cryo-electron microscopy reconstructions. We show that RNA restructuring events coincide with the release of assembly factors and predict that completion of the head domain is required before the Rio1 kinase enters the assembly pathway. Collectively, our results suggest that 40S assembly factors regulate the timely incorporation of ribosomal proteins by delaying specific folding steps in the 3’ major domain of the 20S pre-ribosomal RNA.
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| Overall design |
Three datasets of yeast ribosomal samples subjected to different chemical modifications; 1M7 dataset contains 8 different modified samples and 2 control samples; NAI dataset contains 3 different modified samples and 2 control samples; DMS dataset contains 1 modified sample and 1 control sample. Each sample consists of at least two replicates.
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| |
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| Contributor(s) |
Hector RD, Granneman S |
| Citation(s) |
25200078, 34044851 |
| |
| Submission date |
Dec 02, 2013 |
| Last update date |
Jun 03, 2021 |
| Contact name |
Sander Granneman |
| E-mail(s) |
[email protected]
|
| Organization name |
University of Edinburgh
|
| Department |
Centre for Synthetic and Systems Biology
|
| Lab |
Granneman lab
|
| Street address |
Mayfield Road, Kings Buildings, Waddington building, room 3.06
|
| City |
Edinburgh |
| ZIP/Postal code |
EH9 3JD |
| Country |
United Kingdom |
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| Platforms (1) |
| GPL13821 |
Illumina HiSeq 2000 (Saccharomyces cerevisiae) |
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| Samples (41)
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| Relations |
| BioProject |
PRJNA230385 |
| SRA |
SRP033417 |
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