TY - JOUR
T1 - IFN-β-induced alteration of VSV protein phosphorylation in neuronal cells
AU - D'Agostino, Paul M.
AU - Amenta, Jessica J.
AU - Reiss, Carol Shoshkes
PY - 2009/12/1
Y1 - 2009/12/1
N2 - Vesicular stomatitis virus (VSV) replication is highly sensitive to interferon (IFN)-induced antiviral responses. VSV infection of well-known cell lines pretreated with IFN-β results in a 104-fold reduction in the release of infectious particles, with a concomitant abrogation in viral transcript and/or protein levels. However, in cell lines of neuronal lineage only a threefold reduction in viral transcript and protein levels was observed, despite the same 104-fold reduction in released infectious virions, suggesting an assembly defect. Examination of VSV matrix (M) protein ubiquitination yielded no differences between mock- and IFN-β-treated neuronal cells. Further analysis of potential post-translational modification events, by scintillation and two-dimensional electrophoretic methods, revealed IFN-β-induced alterations in M protein and phosphoprotein (P) phosphorylation. Hypophosphorylated P protein was demonstrated by reduced 32P counts, normalized by 35S-cysteine/methionine incorporation, and by a shift in isoelectric focusing. Hypophosphorylation of VSV P protein was found to occur in neuronal cell lysates, but not within budded virions from the same IFN-β-treated cells. In contrast, hyperphosphorylation of VSV M protein was observed in both cell lysates and viral particles from IFN-β-treated neuronal cells. Hyperphosphorylated M protein was demonstrated by increased 32P counts relative to 35S-cysteine/methionine normalization, and by altered isoelectric focusing in protein populations from cell and viral lysates. Hyperphosphorylated VSV M protein was found to inhibit its association with VSV nucleocapsid, suggesting a possible mechanism for type I IFN-mediated misassembly through disruption of the interactions between ribonucleoprotein cores, and hyperphosphorylated M protein bound to the plasma membrane inner leaflet.
AB - Vesicular stomatitis virus (VSV) replication is highly sensitive to interferon (IFN)-induced antiviral responses. VSV infection of well-known cell lines pretreated with IFN-β results in a 104-fold reduction in the release of infectious particles, with a concomitant abrogation in viral transcript and/or protein levels. However, in cell lines of neuronal lineage only a threefold reduction in viral transcript and protein levels was observed, despite the same 104-fold reduction in released infectious virions, suggesting an assembly defect. Examination of VSV matrix (M) protein ubiquitination yielded no differences between mock- and IFN-β-treated neuronal cells. Further analysis of potential post-translational modification events, by scintillation and two-dimensional electrophoretic methods, revealed IFN-β-induced alterations in M protein and phosphoprotein (P) phosphorylation. Hypophosphorylated P protein was demonstrated by reduced 32P counts, normalized by 35S-cysteine/methionine incorporation, and by a shift in isoelectric focusing. Hypophosphorylation of VSV P protein was found to occur in neuronal cell lysates, but not within budded virions from the same IFN-β-treated cells. In contrast, hyperphosphorylation of VSV M protein was observed in both cell lysates and viral particles from IFN-β-treated neuronal cells. Hyperphosphorylated M protein was demonstrated by increased 32P counts relative to 35S-cysteine/methionine normalization, and by altered isoelectric focusing in protein populations from cell and viral lysates. Hyperphosphorylated VSV M protein was found to inhibit its association with VSV nucleocapsid, suggesting a possible mechanism for type I IFN-mediated misassembly through disruption of the interactions between ribonucleoprotein cores, and hyperphosphorylated M protein bound to the plasma membrane inner leaflet.
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U2 - 10.1089/vim.2009.0057
DO - 10.1089/vim.2009.0057
M3 - Article
C2 - 19951173
AN - SCOPUS:71449089455
SN - 0882-8245
VL - 22
SP - 353
EP - 369
JO - Viral Immunology
JF - Viral Immunology
IS - 6
ER -