Identification and Characterization of a Novel Actin‐associated Protein Recruited to Structures at the Plasma Membrane
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Bibliographic record
Abstract
Listeria monocytogenes is a bacterial pathogen that manipulates the host actin cytoskeleton during various parts of its infectious cycle. This bacterium invades host cells using actin‐dependent internalization mechanisms at the cell membrane and once in the cytosol, it generates an actin‐rich “comet tail” that propels the microbe within the host cytoplasm. To spread from cell‐to‐cell, L. monocytogenes uses very similar comet tail‐like structures to form listeriopods which protrude the host cell membrane during the cell‐to‐cell spreading event. Using a mass spectrometry screen designed to identify actin‐associated proteins that interact at the plasma membrane, we identified a ubiquitin‐conjugating protein (Ubc). To determine the functional role of this protein in cell membrane‐associated actin regulation, we hypothesized that Ubc would play a crucial role in Listeria monocytogenes infections as well as mammalian cell motility in general as actin‐membrane associations are also crucial for whole cell motility. To test this hypothesis, we initially immunolocalized Ubc in Listeria ‐infected Caco2 cells. Ubc localized to the bacterium‐actin interface exclusively when the bacteria were along the cell membrane. To characterize the function of Ubc at these structures, we utilized both a Ubc functional inhibitor on Caco2 cells as well as mouse embryonic fibroblasts that had decreased expression of the Ubc protein. Using both strategies, we observed a decrease in 1) bacterial internalization and 2) the formation of Listeria ‐induced actin structures, compared to infections with wildtype host cells. To determine if Ubc may play a role in whole cell motility, we conducted a wound healing assay on Ubc‐inhibited PtK2 cells and found that wound closure was impeded in Ubc‐inhibited cells after 24 hours as compared to controls. Our data indicates that Ubc is a novel regulator for general cell membrane‐associated actin dynamics and is a crucial factor for efficient bacterial infections. Support or Funding Information This study was funded through NSERC.
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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 it