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. 2026 Oct 22;123(43):e2600336123. doi: 10.1073/pnas.2600336123

Fig. 2.

Thirteen panel figure with graphs, micrographs, and diagrams showing virus protein synthesis and corneal infection data labeled A through M.

Infected cells reduce protein synthesis commensurate with viral attenuation levels. (A) Representative images of puromycin staining for mouse 3T3 fibroblasts infected overnight with MOI 10 of either wild type HSV-1 or vhs null (UL41NHB) or rescued (UL41NHB-R) viruses (all KOS). (B) Quantification of average anti-puromycin fluorescence per cell from (A) All bar plots show the mean ± SD of multiple biological replicates. (C) Representative images of puromycin staining for mouse 3T3 fibroblasts transfected with 200 ng/well of plasmid encoding HSV-1 vhs, or infected with w.t. HSV-1 strain KOS. (D) Quantification of average anti-puromycin fluorescence per cell from (C). All bar plots show the mean ± SD of multiple biological replicates. (E) Representative images of puromycin staining for mouse 3T3 fibroblasts infected overnight with MOI 10 of each of the viral mutant strains (all KOS), or transfected with 200 ng/well of poly dA-dT. (F) Quantification of average anti-puromycin fluorescence per cell from (E). Bars show the mean ± SD of multiple biological replicates. (G) Representative images of puromycin staining for 3T3 fibroblasts infected for varied time with MOI 10 of HSV-1 vp26-mCerulean (KOS). (H) Quantification of average anti-puromycin fluorescence per cell from (G). Bars show the mean ± SD of multiple biological replicates. (I) Cartoon diagram of experimental layout. Mice are infected on debrided corneas with 4 × 106 PFU of HSV-1 vp26-tdTomato. Puromycin incorporation is performed 48 h postinfection, and then whole dissected corneas are imaged using confocal microscopy. (J) Representative maximum intensity image of an infected mouse cornea. The virus channel (pink) is removed from the right image to enable visualization of puromycin staining. (K) Zoom view of the infection site. (L) Close-up view of the indicated cross-section in (J). (M) Violin plots quantifying anti-puromycin fluorescence for VP26-tdTomato+ (infected) or - (uninfected) cells aggregated from 4 infected animals. Inner lines show the median ± quartiles of the data.