Affiliation:
1. Centro de Biologı́a Molecular (CSIC-UAM), Universidad Autónoma de Madrid, Cantoblanco, 28049 Madrid, Spain
Abstract
ABSTRACT
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.
Publisher
American Society for Microbiology
Subject
Virology,Insect Science,Immunology,Microbiology
Cited by
34 articles.
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