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J Biol Chem, Vol. 273, Issue 43, 27960-27967, October 23, 1998

Mutational Analysis of Poliovirus 2A^pro
DISTINCT INHIBITORY FUNCTIONS OF 2A^pro ON TRANSLATION AND TRANSCRIPTION

From the Centro de Biología Molecular (Consejo Superior de Investigaciones Científicas), Universidad Autónoma de Madrid, Canto Blanco, 28049 Madrid, Spain

Transient expression of poliovirus 2A^pro in mammalian cells by means of the recombinant vaccinia virus vT7 expression system leads to drastic inhibition of both cellular and vaccinia virus gene expression (Aldabe, R., Feduchi, E., Novoa, I., and Carrasco, L. (1995) FEBS Lett. 377, 1-5; Aldabe, R., Feduchi, E., Novoa, I., and Carrasco, L. (1995) Biochem. Biophys. Res. Commun. 215, 928-936). To obtain further insights into the molecular basis of this inhibition, a number of 2A^pro variants were generated and expressed in COS-1 cells. The effect of these variants on cellular translation, on vaccinia virus-specific translation, and on transcription of the reporter gene luciferase was analyzed. The ability of the different 2A^pro variants to block cellular translation depends on their capacities to cleave eIF-4G. The blockade exerted 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 polioviruses mutated in 2A^pro were defective in inhibiting luciferase transcription, yet were still able to cleave eIF-4G and block translation. These findings indicate that 2A^pro interferes with cellular gene expression at both the transcriptional and translational levels. Moreover, these two effects probably reflect the inactivation of different host proteins by poliovirus 2A^pro.

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