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| Status |
Public on Sep 14, 2023 |
| Title |
In vivo-like nearest neighbor parameters accurately predict fractional RNA base-pairing in cells |
| Organism |
Escherichia coli str. K-12 substr. MG1655 |
| Experiment type |
Other
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| Summary |
In this study, we conducted a thermodynamic analysis of RNA helix stability in the Eco80 artificial cytoplasm, which mimics in vivo conditions. Eco80 contains 80% of Escherichia coli metabolites, with biological concentrations of metal ions including 2 mM free Mg2+ and 29 mM metabolite-chelated Mg2+. We determined a set of Watson-Crick free energy nearest-neighbor parameters in Eco80 and found that helices are less stable by ∆∆Go37 ~ 1 kcal/mol in comparison to the traditional 1 M NaCl condition. Analysis indicates that Eco80 reduces the stability of three nearest-neighbor pairs. We applied this information to update the nearest-neighbor model using the RNAstructure package. We determined that our in vivo-like corrections have minimal effects on the prediction of RNA secondary structures determined in vitro and in silico. In contrast, in vivo-like corrections markedly improve prediction of fractional RNA base pairing in E. coli, as benchmarked with in vivo RNA chemical probing data using both DMS and EDC. In summary, our thermodynamic and chemical probing analyses of RNA helices in Eco80 indicate that RNA secondary structures are less stable in cells than in artificially stable in vitro buffer conditions such as 1 M NaCl.
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| Overall design |
Structure-seq2 libraries constructed from RNA extracted from Escherichia coli grown to mid-exponential phase at 37 °C in LB media. Control samples, DMS-treated samples, and EDC-treated samples were prepared as biological replicates. Libraries were sequenced on an Illumina NextSeq platform in single-end mode.
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| Contributor(s) |
Sieg JP, Jolley EA, Hout MJ, Babitzke P, Bevilacqua PC |
| Citation(s) |
37855684 |
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| Submission date |
May 26, 2023 |
| Last update date |
Dec 14, 2023 |
| Contact name |
Elizabeth Jolley |
| E-mail(s) |
eaj19@psu.edu
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| Phone |
8148633109
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| Organization name |
Pennsylvania State University
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| Lab |
Bevilacqua
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| Street address |
239 Chemistry Bldg, Pennsylvania State University
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| City |
University Park |
| State/province |
PA |
| ZIP/Postal code |
16802 |
| Country |
USA |
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| Platforms (1) |
| GPL33080 |
NextSeq 2000 (Escherichia coli str. K-12 substr. MG1655) |
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| Samples (9)
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| GSM7431300 |
WT, control, rep 2 |
| GSM7431301 |
WT, DMS, rep 2 |
| GSM7431302 |
WT, EDC, rep 2 |
| GSM7431303 |
WT, control, rep 3 |
| GSM7431304 |
WT, DMS, rep 3 |
| GSM7431305 |
WT, EDC, rep 3 |
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| Relations |
| BioProject |
PRJNA976738 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSE233607_RAW.tar |
42.3 Mb |
(http)(custom) |
TAR (of TSV, TXT) |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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