Prophage terminase with tRNase activity sensitizes S. enterica to oxidative stress

Phage viruses shape the evolution and virulence of their bacterial hosts. The Salmonella enterica genome encodes several stress-inducible prophages. The Gifsy-1 prophage terminase protein, whose canonical function is to process phage DNA for packaging in the virus head, unexpectedly acts as a tRNase under oxidative stress, cleaving the anticodon loop of tRNALeu. The ensuing RNA fragmentation compromises bacterial translation, intracellular survival, and recovery from oxidative stress in the vertebrate host. S. enterica adapts to this tRNA fragmentation by transcribing the RNA repair Rtc system. The counterintuitive translational arrest provided by tRNA cleavage may subvert prophage mobilization and give the host an opportunity for repair as a way of maintaining bacterial genome integrity and ultimately survival in animals., Assessment of de novo protein synthesis. Protein translation was determined by Western blot version of SUnSET as described previously with minor modifications (33). Briefly, Salmonella stains were grown in MOPS-GLC minimal media pH-7.2 at 37 ºC to an OD600 of ~ 0.25 in a shaking incubator. The cells were treated with 400 mM H2O2 for 10 min, followed by an additional 40 min incubation in the presence of 250 mg/ml puromycin. Cytoplasmic proteins were extracted and quantified by the BCA method. 30 mg of protein samples were separated on 10% SDS-polyacrylamide gels. The specimens were transferred onto nitrocellulose membranes and immunoblotted using anti-puromycin antibody (Millipore). Protein synthesis was quantified from the immunoblots using the entire molecular weight range of puromycin-incorporated in each lane. The relative puromycin-incorporated levels were quantified with the ImageJ software (NIH). Ponceau stained membrane was used to make sure protein loading was comparable across ..., , # Prophage terminase with tRNAse activity sensitizes Salmonella to oxidative stress

[https://doi.org/10.5061/dryad.xpnvx0knt](https://doi.org/10.5061/dryad.xpnvx0knt)

This dataset contains unprocessed images and raw data generated in the study. Interpretation of data should be made in conjunction with the main research article.

## Description of the data and file structure

Raw data for each figure is provided in Excel file with each sheet corresponding to each panel in the figure.

The uncropped northern and western blot images and histopathology images are uploaded as tif/jpg images in the linked Zenodo repository.

#### **Fig 1.**

(A, D) Immunoblots with Ponceau-S-stained controls and (B, E) a corresponding densitometric analysis examining puromycin incorporation by *S. enterica* 30 minutes after H2O2 treatment. Densitometry was normalized against Ponceau S-stained lanes using ImageJ.

(C) Killing of *S. enterica* by H2O2 in PBS after 30 minutes of treatment.  

(F) Differentiall...