MUTATIONAL ANALYSIS OF POLIOVIRUS 2A(PRO) - DISTINCT INHIBITORY FUNCTIONS OF 2A(PRO) ON TRANSLATION AND TRANSCRIPTION

Transient expression of poliovirus 2A(pro) in mammalian cells by means of the recombinant vaccinia virus vT7 expression system leads to dras tic inhibition of both cellular and vaccinia virus gene expression (Al dabe, R., Feduchi, E., Novoa, I., and Carrasco, L. (1995) FEBS Lett. 3 77, 1-5; Aldabe, R., Feduchi, E., Novoa, I., and Carrasco, L. (1995) B iochem. Biophys. Res. Commun. 215, 928-936). To obtain further insight s into the molecular basis of this inhibition, a number of 2A(pro) var iants were generated and expressed in COS-1 cells. The effect of these variants on cellular translation, on vaccinia virus-specific translat ion, and on transcription of the reporter gene luciferase was analyzed . The ability of the different 2A(pro) variants to block cellular tran slation depends on their capacities to cleave eIF-4G. The blockade exe rted by 2A(pro) on transcription of the luciferase gene reinforces the notion that this protease is a potent inhibitor of RNA polymerase II- mediated transcription. Some of the 2A(pro) variants tested failed to block luciferase transcription, despite the fact that eIF-4G cleavage and inhibition of translation were observed. Two reconstituted poliovi ruses mutated in 2A(pro) were defective in inhibiting luciferase trans cription, yet were still able to cleave eIF-4G and block translation. These findings indicate that 2A(pro) interferes with cellular gene exp ression at both the transcriptional and translational levels. Moreover , these two effects probably reflect the inactivation of different hos t proteins by poliovirus 2A(pro).