Fig. 9. Effect of nocodazole on SEAP secretion in uninfected and poliovirus-infected cells. COS-7 cells were transfected with a plasmid, pIRES-SEAP, that expressed (A) SEAP protein under the translational control of the IRES element from encephalomyocarditis virus (EMCV) to ensure effective translation in poliovirus-infected cells. (B) Transfected COS-7 cells were incubated for 16 hours at 37°C with or without 2 µg/ml BFA as indicated. Aliquots from the supernatants were sampled every hour from both cultures and assayed for SEAP activity. (C) Transfected COS-7 cells were either incubated with serum-free medium with or without 5 µg/ml nocodazole for 1.5 hours The medium was then replaced with fresh medium with or without nocodazole and aliquots taken every hour to determine SEAP activity. (D) Transfected COS-7 cells were mock-infected or infected with poliovirus at an MOI of 10 PFU/cell. SEAP activity was assayed in both the supernatants and the cells of mock-infected and infected cells at the time points indicated. The percentage of SEAP activity in infected cells compared with that in uninfected cells is plotted for secreted and intracellular SEAP as a function of time post infection. Experiments were performed in triplicate and the standard error is shown. (E) pIRES-SEAP-transfected cells were treated for 1.5 hours with serum-free medium with or without 5 µg/ml nocodazole and either mock-infected or infected with poliovirus at an MOI of 10 PFU/cell. Aliquots of medium were sampled every hour and assayed for SEAP activity. The percentage of extracellular and intracellular SEAP activity in infected cells compared with uninfected cells as a function of time after infection in the presence of nocodazole is shown.