Twitching motility suppressors reveal a role for FimX in type IV pilus extension dynamics
Bibliographic record
Abstract
Abstract In Pseudomonas aeruginosa, retractable protein filaments called type IV pili (T4P) facilitate surface adherence, sensing, and directional movement known as twitching motility. T4P are necessary for the bacteria to engage in surface-associated behaviors, including establishing acute infections. Pilus extension is driven by the hexameric ATPase, PilB, at the base of the T4P nanomachine in coordination with various protein regulatory effectors. The cyclic-di-GMP binding protein, FimX, works with PilB to mediate normal extension processes, though how this effector controls pilus assembly remains unclear. To explore the role of FimX in T4P function, we leveraged the significant Δ fimX twitching motility deficit to screen for mutants capable of overcoming this phenotype. We identified suppressor mutations that increase twitching in Δ fimX background, mapping primarily to cyclic-AMP homeostatic machinery or to PilB, the FimX target. Distinct suppressor mutations in PilB increased ATP hydrolysis in vitro and this activity was subject to modulation by FimX. Using microscopy to monitor the extension dynamics of fluorescently labelled T4P, we showed that Δ fimX mutants produce slow-to-extend, short pili, a phenotype that is rescued by mutations enhancing PilB ATP hydrolysis and/or re-introduction of FimX. Together, these data implicate FimX as a regulator of PilB enzymatic function, potentially enabling P. aeruginosa to fine-tune pilus extension dynamics in response to environmental cues. Summary Type IV pili enable Pseudomonas aeruginosa to attach to surfaces, move (twitch), and form biofilms. Pilus extension is powered by the motor protein PilB, which is regulated by other factors, including FimX, a protein that binds cyclic-di-GMP. Although FimX is important for twitching, how it influences PilB was unclear. We deleted fimX , which severely reduces motility, and searched for mutants that regained movement. We identified two types: some had mutations in PilB that increased its ATPase activity, allowing it to function without FimX, while others affected the cyclic-AMP signaling pathway and increased overall production of pilus components, showing that motility can also be improved through changes in quantity versus quality. Our results suggest that FimX normally fine-tunes PilB enzymatic activity, enabling dynamic control of pilus extension in response to surface signals. This work helps explain how P. aeruginosa adapts to different environments, a process crucial for infection and biofilm development.
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How this classification was reachedexpand
Full frame machine prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. The Gemma side is a direct model label for every work in the frame, read from the title-only record. The Codex side is a classifier learned from the 10,348 direct Codex labels and calibrated to design-weighted sample rates; fields without enough sample support carry no Codex call. Candidate is the union of the two sides; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels.
Distilled classifier scores by category (both heads)
| 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 | 0.001 |
| Insufficient payload (model declined to judge) | 0.001 | 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 source (direct Gemma or distilled Codex), 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".