A Stable HeLa Cell Line That Inducibly Expresses Poliovirus 2Apro: Effects on Cellular and Viral Gene Expression

摘要

A HeLa cell clone (2A7d) that inducibly expresses the gene for poliovirus protease 2A (2A^pro) under the control of tetracycline has been obtained. Synthesis of 2A^pro induces severe morphological changes in 2A7d cells. One day after tetracycline removal, cells round up and a few hours later die. Poliovirus 2A^pro cleaves both forms of initiation factor eIF4G, causing extensive inhibition of capped-mRNA translation a few hours after protease induction. Methoxysuccinyl-Ala-Ala-Pro-Val-chloromethylketone, a selective inhibitor of 2A^pro, prevents both eIF4G cleavage and inhibition of translation but not cellular death. Expression of 2A^pro still allows both the replication of poliovirus and the translation of mRNAs containing a picornavirus leader sequence, while vaccinia virus replication is drastically inhibited. Translation of transfected capped mRNA is blocked in 2A7d-On cells, while luciferase synthesis from a mRNA bearing a picornavirus internal ribosome entry site (IRES) sequence is enhanced by the presence of 2A^pro. Moreover, synthesis of 2A^pro in 2A7d cells complements the translational defect of a poliovirus 2A^pro-defective variant. These results show that poliovirus 2A^pro expression mimics some phenotypical characteristics of poliovirus-infected cells, such as cell rounding, inhibition of protein synthesis and enhancement of IRES-driven translation. This cell line constitutes a useful tool to further analyze 2A^pro functions, to complement poliovirus 2A^pro mutants, and to test antiviral compounds.