Alphavirus nsP3 ADP-ribosylhydrolase Activity Disrupts Stress Granule Formation

Aravinth Kumar Jayabalan; Diane E. Griffin; Anthony K. L. Leung

The copyright holder for this preprint (which was not peer-reviewed) is the author/funder. . https://doi.org/10.1101/629881 doi: bioRxiv preprint Page 16 protein nsP3 through its ability to enzymatically remove ADP-ribosylation. Expression of the wild-type nsP3 MD alone is sufficient to suppress SG formation. The requirement for ADP-ribosylhydrolase activity was demonstrated by the loss of ability to suppress SG formation by five different mutants with impaired enzyme activity. Though it is possible that activities other than ADP-ribosylhydrolase possessed by the nsP3 MD may be responsible for suppressing SG formation, the fact that mutations at different sites in the MD result in the same phenotype make such a scenario less likely. Importantly, the expression of WT nsP3, but not the ADP-ribosylhydrolase-deficient mutant nsP3s, suppresses the formation of SGs in response to multiple types of stress including oxidative stress (arsenite), mitochondrial stress (clotrimazole) and ER stress (thapsigargin). Suppression was observed both by ectopic expression of nsP3 through transfection and natively in the context of CHIKV infection. Given that the MD and its ADP-ribosylhydrolase activity are conserved across all macrodomain-containing RNA viruses [66, 67] , including coronaviruses, it is possible that these viruses suppress SG formation through a common mechanism by removing ADP-ribosylation.