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J Virol, May 1998, p. 3560-3570, Vol. 72, No. 5
Centro de Biología Molecular
(CSIC-UAM), Universidad Autónoma de Madrid, Cantoblanco, 28049 Madrid, Spain
Received 22 September 1997/Accepted 20 January 1998
The expression of poliovirus 2BC protein in yeast and mammalian
cells leads to a number of metabolic and morphological alterations, such as growth inhibition, intracellular membrane proliferation, blockade of the exocytic pathway, and enhanced membrane permeability. Yeast cells that express poliovirus 2BC in an inducible manner were
used to identify the regions of 2BC implicated in the modifications of
these cellular functions. Several 2BC deletion mutants were generated
to define the minimal portion of 2BC required to alter these
activities. Additional deletion mutants that were obtained by random
mutagenesis followed by selection in yeast cells provided new insights
into the structure and mechanism of action of 2BC. The activity
responsible for membrane proliferation is located in 2C, while the
activities responsible for membrane permeabilization and inhibition of
the exocytic pathway are located in 2B. Several regions of 2B and 2C
required for the different functions of 2BC were identified. Thus, the
integrity of the N termini of both 2B and 2C is necessary for
2BC-induced cytotoxicity. It is also possible to separate the different
cellular alterations provoked by 2BC by the use of several 2BC
variants. Deletion of amino acids 52 to 65 in 2B generates a 2BC
deletion variant, 2bC
0022-538X/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.
Identification of Regions of Poliovirus 2BC Protein
That Are Involved in Cytotoxicity
AvrII, that still blocks yeast growth but is
unable to enhance membrane permeability or to inhibit the exocytic
pathway. On the other hand, 2Bc128*.32b and 2Bc128*.3c, which contain
only 73 and 77 amino acids of 2B, interfere with yeast division and
enhance membrane permeability but affect the exocytic pathway only
weakly and do not induce membrane proliferation. Our findings indicate
that Saccharomyces cerevisiae represents a useful model
system to analyze the functions of poliovirus 2BC and show the
feasibility of separating the activities assigned to this protein.
*
Corresponding author. Mailing address: Centro de
Biología Molecular (CSIC-UAM), Universidad Autónoma de
Madrid, Cantoblanco, 28049 Madrid, Spain. Phone: 34-1-397 8450. Fax:
34-1-397 4799. E-mail: abarco{at}trasto.cbm.uam.es.
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