Fig 4. The asTORi PP242 suppresses host and viral protein synthesis in normal cells but sustains HSV1-dICP0 protein synthesis in cancer cells.

(A) 4T1, NMuMG and NT2196 cells were infected in triplicates with GFP-expressing HSV1-dICP0 (0.1 MOI) in the presence of DMSO or PP242 (2μM), pretreated for 30 min prior to infection. Global protein synthesis was assessed by a 30 min pulse [³⁵S]methionine incorporation into newly synthesized proteins at 2 hours and 24 hours post-infection. Triplicate cell lysates were separated on SDS-PAGE and changes in protein synthesis were revealed by autoradiography (top), [³⁵S]methionine incorporation was measured by TCA precipitation and quantified on scintillation counter (middle–results presented are normalized to DMSO control set to 100% ± SD (n = 3)), and 4E-BP1 phosphorylation status as well as the levels of HSV1 proteins were assessed by Western blot (bottom). (B) Polysome profile analysis of 4T1, NMuMG and NT2196 cells treated and infected with HSV1-dICP0 as above for 24 hours.