Viruses, Vol. 12, Pages 1021: Venezuelan Equine Encephalitis Virus nsP3 Phosphorylation Can Be Mediated by IKK β Kinase Activity and Abrogation of Phosphorylation Inhibits Negative-Strand Synthesis

Viruses, Vol. 12, Pages 1021: Venezuelan Equine Encephalitis Virus nsP3 Phosphorylation Can Be Mediated by IKKβ Kinase Activity and Abrogation of Phosphorylation Inhibits Negative-Strand Synthesis Viruses doi: 10.3390/v12091021 Authors: Bakovic Bhalla Kortchak Sun Zhou Ahmed Risner Klimstra Narayanan Venezuelan equine encephalitis virus (VEEV), a mosquito transmitted alphavirus of the Togaviridae family, can cause a highly inflammatory and encephalitic disease upon infection. Although a category B select agent, no FDA-approved vaccines or therapeutics against VEEV currently exist. We previously demonstrated NF-κB activation and macromolecular reorganization of the IKK complex upon VEEV infection in vitro, with IKKβ inhibition reducing viral replication. Mass spectrometry and confocal microscopy revealed an interaction between IKKβ and VEEV non-structural protein 3 (nsP3). Here, using western blotting, a cell-free kinase activity assay, and mass spectrometry, we demonstrate that IKKβ kinase activity can directly phosphorylate VEEV nsP3 at sites 204/5, 142, and 134/5. Alanine substitution mutations at sites 204/5, 142, or 134/5 reduced VEEV replication by >30-100,000-fold corresponding to a severe decrease in negative-strand synthesis. Serial passaging rescued viral replication and negative-strand synthesis, and sequencing of revertant viruses revealed reversion to the wild-type TC...
Source: Viruses - Category: Virology Authors: Tags: Article Source Type: research