FIG. 6.

Inhibition of protein synthesis by MV-N in vivo. (A to D) 293-MVN cells or 293 cells were plated in 24-well plates and infected with a recombinant adenovirus, AxCANCre, expressing Cre recombinase at an MOI of 10, 20, or 50. (A) At 24 h postinfection, switching expression of MV-N in 293-MVN cells was detected by Western blotting. (B) At 24 h postinfection, a reporter plasmid encoding the luciferase gene was transfected and incubated for 24 h. Cells were harvested, an aliquot was lysed, and luciferase activity was measured by using a luminometer. Another aliquot was harvested, and the total RNA was analyzed for luciferase transcripts using one-step real-time RT-PCR. The luciferase activity and mRNA levels in 293-MVN cells are expressed relative to those in 293 cells. The results shown represent means of three experiments. (C) At 24 h postinfection, cells were labeled with [³⁵S]methionine for 24 h. An aliquot of the cells was lysed, and the level of radioactivity incorporated in all proteins was measured by using a scintillation counter. An aliquot of the cells was harvested, and the total RNA was analyzed for transcripts of the housekeeping gene, GAPDH, using one-step real-time RT-PCR. The levels of radioactivity and GAPDH mRNA in 293-MVN cells are expressed relative to those in 293 cells. The results shown represent means of three experiments. (D) At 48 h postinfection, cells were lysed and endogenous GAPDH was detected by Western blotting.