Acrylamide inhibits vaccinia virus through vimentin‐independent anti‐viral granule formation

Abstract

The replication and assembly of vaccinia virus (VACV), the prototypic poxvirus, occurs exclusively in the cytoplasm of host cells. While the role of cellular cytoskeletal components in these processes remains poorly understood, vimentin-a type III intermediate filament-has been shown to associate with viral replication sites and to be incorporated into mature VACV virions. Here, we employed chemical and genetic approaches to further investigate the role of vimentin during the VACV lifecycle. The collapse of vimentin filaments, using acrylamide, was found to inhibit VACV infection at the level of genome replication, intermediate-and late-gene expression. However, we found that CRISPR-mediated knockout of vimentin did not impact VACV replication. Combining these tools, we demonstrate that acrylamide treatment results in the formation of anti-viral granules (AVGs) known to mediate translational inhibition of many viruses. We conclude that vimentin is dispensable for poxvirus replication and assembly and that acrylamide, as a potent inducer of AVGs during VACV infection, serves to bolster cell's anti-viral response to poxvirus infection.

Poxviruses package viral redox proteins in lateral bodies and modulate the host oxidative response

“…That RPL17, and therefore presumably intact ribosomes, were found in the final pellet of uninfected “virion purifications”, future assignment of ribosomal proteins as virions components should be verified by super-resolution imaging modalities. In addition to RPL17 and Tomm20, we recently showed that vimentin—which localizes to viral replication sites and is packaged into virions [ 67 , 81 ]–also biochemically fractionates with Core-LBs, and like other LB candidates is trypsin sensitive [ 82 ]. Strikingly, 6 of the host proteins were found to be more abundant in LBs than F17 raising the possibility of active LB incorporation.…”

Coding-Complete Genome Sequences of Copenhagen and Copenhagen-Derived vP811 Strains of Vaccinia Virus Isolated from Cell Culture

“…Laboratory Cop and vP811 stocks were obtained from the University of Alberta and stored in 1 mM Tris buffer, pH 9, at −80°C. Confluent monolayers of BSC-40 cells (ATCC CRL-2761) were infected at a multiplicity of infection of 0.02, and genomic viral DNA was obtained by phenol-chloroform extraction 3 days postinfection as described previously ( 4 , 5 ).…”

Acrylamide Inhibits Vaccinia Virus Through Vimentin-independent Anti-Viral Granule Formation

“…Samples were immunoblotted for A10 (core), F17 (LB) and vimentin. LBs, lateral bodies; VACV, vaccinia virus pre-or post-genome replication, we infected HeLa cells with recombinant viruses that express EGFP under the control of early (WR E-EGFP) or late (WR L-EGFP) viral promoters and analysed them for EGFP expression by flow cytometry (Yakimovich et al, 2017). When plotted as a histogram (cell number vs. fluorescence intensity), no defect in either the number of infected cells or intensity of EGFP expression was seen with WR E-EGFP virus in the presence of acrylamide (Figure 2c; left panel).…”