In vitro characterization of 6S RNA release-defective mutants uncovers features of pRNA-dependent release from RNA polymerase in <i>E. coli</i>
Bibliographic record
Abstract
6S RNA is a noncoding RNA that inhibits bacterial transcription by sequestering RNA polymerase holoenzyme (Eσ(70)) in low-nutrient conditions. This transcriptional block can be relieved by the synthesis of a short product RNA (pRNA) using the 6S RNA as a template. Here, we selected a range of 6S RNA release-defective mutants from a high diversity in vitro pool. Studying the release-defective variant R9-33 uncovered complex interactions between three regions of the 6S RNA. As expected, mutating the transcriptional start site (TSS) slowed and partially inhibited release. Surprisingly, additional mutations near the TSS were found that rescued this effect. Likewise, three mutations in the top strand of the large open bubble (LOB) could considerably slow release but were rescued by the addition of upstream mutations found between a highly conserved "-35" motif and the LOB. Combining the three top strand LOB mutations with mutations near the TSS, however, was particularly effective at preventing release, and this effect could be further enhanced by inclusion of the upstream mutations. Overexpressing R9-33 and a series of milder release-defective mutants in Escherichia coli resulted in a delayed entry into exponential phase together with a decrease in cell survival that correlated well with the severity of the in vitro phenotypes. The complex crosstalk observed between distinct regions of the 6S RNA supports a scrunching type model of 6S RNA release, where at least three regions of the 6S RNA must interact with Eσ(70) in a cooperative manner so as to ensure effective pRNA-dependent release.
Fetched live from OpenAlex and de-inverted. Abstracts are not stored in this database: the inverted indexes are 8.6 GB of the frame’s 9.3 GB of text, and the host has 13 GB free.
How this classification was reachedexpand
Full frame distilled prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. Learned from the 10,348 direct Codex labels and 10,348 direct Gemma labels. Candidate is the union of thresholded teacher heads; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels or direct frontier model labels.
Codex and Gemma teacher scores by category
| Category | Codex | Gemma |
|---|---|---|
| Metaresearch | 0.000 | 0.000 |
| Meta-epidemiology (narrow) | 0.000 | 0.000 |
| Meta-epidemiology (broad) | 0.000 | 0.000 |
| Bibliometrics | 0.000 | 0.000 |
| Science and technology studies | 0.000 | 0.000 |
| Scholarly communication | 0.000 | 0.000 |
| Open science | 0.000 | 0.000 |
| Research integrity | 0.000 | 0.000 |
| Insufficient payload (model declined to judge) | 0.000 | 0.000 |
Machine scores (provisional)
The two teacher heads of the student model, read on this work. A score orders the frame for review; it never asserts a category, and the validation status ships verbatim with every row.
Baseline scores from an immature model (maturity gate not passed, 7 training rounds). Scores rank; they never assert a category.
score_only:v0-immature-baseline · verbatim from the scoring run: score_only means the number may rank works, and no category label ships from itClassification
machine, unvalidatedMachine predicted; a candidate call from one teacher head, not a consensus.
How this classification was reached, model by model and score by score, is at the end of the page under "How this classification was reached".