Opposing Roles of Double-Stranded RNA Effector Pathways and Viral Defense Proteins Revealed with CRISPR-Cas9 Knockout Cell Lines and Vaccinia Virus Mutants.
Ruikang LiuBernard MossPublished in: Journal of virology (2016)
Viruses have evolved ways of preventing or counteracting the cascade of antiviral responses that double-stranded RNA (dsRNA) triggers in host cells. We showed that the dsRNA produced in excess in cells infected with a vaccinia virus (VACV) decapping enzyme mutant and by wild-type virus colocalized with the viral E3 protein in cytoplasmic viral factories. Novel human cell lines defective in either or both protein kinase R and RNase L dsRNA effector pathways and/or the cellular 5' exonuclease Xrn1 were prepared by CRISPR-Cas9 gene editing. Inactivation of both pathways was necessary and sufficient to allow full replication of the E3 mutant and reverse the defect cause by inactivation of Xrn1, whereas the decapping enzyme mutant still exhibited defects in gene expression. The study provided new insights into functions of the VACV proteins, and the well-characterized panel of CRISPR-Cas9-modified human cell lines should have broad applicability for studying innate dsRNA pathways.
Keyphrases
- wild type
- crispr cas
- genome editing
- induced apoptosis
- gene expression
- endothelial cells
- sars cov
- cell cycle arrest
- binding protein
- immune response
- protein kinase
- induced pluripotent stem cells
- nucleic acid
- endoplasmic reticulum stress
- dna methylation
- regulatory t cells
- single cell
- oxidative stress
- type iii
- innate immune
- soft tissue
- genetic diversity