FIG. 5.

Effects of SB203580 on EMC viral transcription and protein synthesis. L929 cells were infected with EMC virus (MOI = 5), and fresh medium with (solid circles) or without (open circles) SB203580 (40 μM) was added 8 h later. (A) Total RNA was isolated after 0, 1, 3, and 5 h of SB203580 treatment, and the RPA was performed with an EMC virus-specific probe. β-Actin mRNA was measured as an internal standard (upper panel). Relative EMC viral RNA synthesis was measured as a ratio of β-actin mRNA (lower panel). (B) Cell lysates were prepared after 0, 1, 3, and 5 h of SB203580 treatment, and Western blotting was performed with polyclonal anti-EMC virus antibodies for VP1 (EMCVP1) or anti-β-actin antibodies (upper panel). (C) Cells were labeled with [³⁵S]methionine for 30 min after 0, 1, 3, and 5 h of SB203580 treatment, and cell lysates were immunoprecipitated with polyclonal anti-EMC virus antibodies for VP1 (EMC VP1) or anti-β-actin antibodies. The samples were separated by SDS-PAGE and analyzed by autoradiography (upper panel). Relative EMC protein synthesis was measured as ratio of β-actin protein (B and C, lower panel). Nonsaturated film exposures were selected, scanned, and imported into the UTHSCASA Image Tool for quantification.