A human virus protein, poliovirus protein 2BC, induces membrane proliferation and blocks the exocytic pathway in the yeast Saccharomyces cerevisiae.

Abstract

Inducible synthesis of poliovirus protein 2BC in Saccharomyces cerevisiae arrests cell growth in the G2 phase of the cell cycle, while no effects are observed upon expression of poliovirus genes 2B or 2C, either individually or in combination. Expression of 2BC induces a number of morphological modifications in yeast cells, one of the most striking being the proliferation of small membranous vesicles that fill most of the cytoplasm. These vesicles are morphologically similar to the cytopathic vacuoles that proliferate during the infection of human cells by poliovirus. The transport and processing of several yeast proteins, including vacuolar carboxypeptidase Y, aminopeptidase I or yeast alpha‐mating factor, is hampered upon expression of poliovirus 2BC, suggesting that transport of proteins through the Golgi apparatus is impaired by this viral protein. Finally, a number of 2BC variants were generated and the effects of their expression on yeast growth, cellular morphology and protein processing were analyzed. 2BC variants defective in the NTPase activity were still able to interfere with yeast growth and the exocytic system, while deletion of 30 amino acids at the N‐terminus of 2BC impairs its function. These findings lend support to the idea that 2BC, but not 2B or 2C, is the protein responsible for vesicle proliferation in poliovirus‐infected cells. In addition, the activity of a human virus protein in yeast cells opens new avenues to investigate the exact location at which poliovirus 2BC interferes with the vesicular system and to test the action of other animal virus proteins potentially involved in modifying the vesicular system in mammalian cells.

Poliovirus infection blocks ERGIC-to-Golgi trafficking and induces microtubule-dependent disruption of the Golgi complex

“…Analysis of the effects of individual viral proteins may facilitate understanding the effects of viral infection, although their combined action during infection may not be successfully mimicked. Further investigation will be required to link our findings that poliovirus infection blocks the ERGIC-to-Golgi step with the effects observed upon ectopic expression of various viral proteins that affect protein secretion (Barco and Carrasco, 1995;Cho et al, 1994;Doedens et al, 1997;Doedens and Kirkegaard, 1995;Sandoval and Carrasco, 1997).…”

“…Protein 3A employs a Cterminal hydrophobic domain that confers all the biophysical properties expected in an integral membrane protein (Wessels et al, 2006). Proteins 2BC and 3A have also been shown to exert potent effects on intracellular membranes when expressed individually in cells (Cho et al, 1994;Barco and Carrasco, 1995;Doedens et al, 1997;Suhy et al, 2000). For example, the expression of poliovirus 3A protein inhibits protein secretion and colocalizes with ER markers (Doedens and Kirkegaard, 1995).…”

Viroporins: structure, function and potential as antiviral targets

“…2BC, 2B, 2C) were initially shown to modulate both membrane permeability (Aldabe et al, 1996;Barco & Carrasco, 1995) and membrane trafficking (Doedens & Kirkegaard, 1995), yet 2B is now commonly accepted as the principle mediator of such behaviour. 2B is a class 2 viroporin with two helical TMDs separated by a stretch of highly polar residues.…”

Poliovirus 2Apro expression inhibits growth of yeast cells

“…TCA precipitation was used to determine incorporation levels of radioactive precursors. Western blot analysis was carried out as described [19].…”