Myxoma Virus Protein M029 Is a Dual Function Immunomodulator that Inhibits PKR and Also Conscripts RHA/DHX9 to Promote Expanded Host Tropism and Viral Replication
Figure 4
M029 is essential for MYXV replication in cell lines originated from diverse species.
Single step growth curves of MYXV infection in A) BSC40, B) NIH3T3 and C) BHK21 cells. The indicated cells were infected with vMyx-GFP, vMyx-M029KO or vMyx-M029ID at an MOI of 5, and then cells were collected at 0, 6, 12, 24, 36, and 48 h p.i. virus titers were determined in triplicate following serial dilution onto RK13-E3 cells. Data are representative of two independent experiments. D) Fluorescence microscope images of infected BSC40 (top panels), NIH3T3 (middle panels) and BHK21 (bottom panels) cells. The images were taken using an inverted fluorescence microscope using a lens with ×10 magnification at 48 h p.i. The cells were infected with vMyx-GFP (left column), vMyx-M029KO (middle column) and vMyx-M029ID (right column). E) Expression of viral early (M-T7) and late (Serp-1) proteins in the infected BHK21 and NIH3T3 cells. BHK21 and NIH3T3 cells were left uninfected (M) (lanes 1 and 8) or were infected with vMyxGFP (lanes 2 to 7) or vMyx-M029ID (lanes 9 to 14) at an MOI of 3. Cells were collected at 2 (lanes 2 and 9), 4 (lanes 3 and 10), 8 (lanes 4 and 11), 12 (lanes 5 and 12), 24 (lanes 6 and 13) and 24a in the presence of AraC (lanes 7 and 14) h p.i. The membranes were first probed with anti-Serp-1 antibody, stripped and probed for M-T7 and actin (loading control).