Fig. 3. CSDE1 regulates viral M RNA levels.

Cells (HepB3 or HepB3-CSDE1^C-T; B16 or B16-CSDE1^C-T) were infected with VSV viruses (VSV-IFNβ or VSV-IFNβ-IGR P/M^C-U) at an MOI of 3. Lanes: 1, Hep3B/VSV-hIFNβ; 2, Hep3B/VSV-hIFNβ-IGR P/M^C-U; 3, Hep3B-CSDE1^C-T/VSV-hIFNβ; 4, Hep3B-CSDE1^C-T/VSV-hIFNβ-IGR P/M^C-U; 5, B16/VSV-mIFNβ; 6, B16/VSV-mIFNβ-IGR P/M^C-U; 7, B16-CSDE1^C-T/VSV-mIFNβ; and 8, B16-CSDE1^C-T/ VSV-mIFNβ-IGR P/M^C-U. Six hours later, RNA was isolated and converted into cDNA. qrtPCR was used to assess levels of A viral P RNA (P-specific primers P1 and P2), B viral M RNA (M-specific primers M1 and M2), or C viral P–M RNA (P/M IGR-M-specific primers IGR1 and M2). Representative of three separate experiments. D–F Levels of G, L, and G–L RNA were measured as in A–C using primers G1 and 2, L1 and 2, and G1–L2, respectively. Representative of two separate experiments. Levels of each RNA species were normalized to expression in Hep3B infected with VSV-hIFNβ or to B16 infected with VSV-mIFNβ. G Hep3B parental cells (Hep3B) (Lanes 1–3) or Hep3B cells engineered to overexpress CSDE1^wt (Hep3B-CSDE1^wt) (lanes 4 and 5) or mutated CSDE1^P5S (Hep3B-CSDE1^C-T) (lanes 6 and 8) proteins, or Hep3B that had been selected in vitro for resistance to VSV-IFNβ oncolysis for 21 days (Hep3B-VSV-IFNβ-ESC) (lanes 7 and 9), were infected with wild-type VSV-IFNβ (lanes 2 and 5–7) or VSV-IFNβ-IGR P/M^C-U (lanes 3, 4, 8, and 9) at an MOI of 3. Cells were collected at either 6 or 18 h post infection and levels of viral M protein were measured by western blotting. Representative of two separate experiments means ± SD of three biological replicates are shown (A–F). P-values were determined using a one-way ANOVA (A–F) on the raw data. Statistical significance was set at p < 0.05, ns > 0.05. *p < 0.05, **p < 0.1, ***p < 0.001. Source data are provided as a Source Data file.