bioRxiv2020
Yellow fever virus (YFV) is an RNA virus primarily targeting the liver. Severe YF cases are responsible for hemorrhagic fever, plausibly precipitated by excessive pro-inflammatory cytokine response. Pathogen recognition receptors (PRRs), such as the cytoplasmic RIG-I-like receptors (RLRs), and the viral RNA sensor PKR are known to initiate a pro-inflammatory response upon recognition of viral genomes. Here, we sought to reveal the main determinants responsible for the acute cytokine expression occurring in human hepatocytes following YFV infection. Using a RIG-I-defective human hepatoma cell line, we found that RIG-I largely contributes to cytokine secretion upon YFV infection. In infected RIG-I-proficient hepatoma cells, RIG-I was localized in stress granules. These granules are large aggregates of stalled translation preinitiation complexes known to concentrate RLRs and PKR, and are so far recognized as hubs orchestrating RNA virus sensing. Using PKR-deficient hepatoma cells, we found that PKR contributes to both stress granule formation and cytokine induction upon YFV infection. However, stress granules disruption did not affect the cytokine response to YFV infection, as assessed by siRNA-knockdown-mediated inhibition of stress granule assembly. Finally, no viral RNA was detected in stress granules using a fluorescence in situ hybridization approach coupled with immunofluorescence. Our findings suggest that both RIG-I and PKR mediate pro-inflammatory cytokine induction in YFV-infected hepatocytes, in a stress granule-independent manner. Therefore, by showing the uncoupling of the early cytokine response from the stress granules formation, our model challenges the current view by which stress granules are required for the mounting of the acute antiviral response. ImportanceYellow fever is a mosquito-borne acute hemorrhagic disease caused by yellow fever virus (YFV). The mechanisms responsible for its pathogenesis remain largely unknown, although increased inflammation has been linked to worsened outcome. YFV targets the liver, where it primarily infects hepatocytes. We found that two RNA-sensing proteins, RIG-I and PKR, participate in the induction of pro-inflammatory mediators in human hepatocytes infected with YFV. We show that YFV infection promotes the formation of cytoplasmic structures, termed stress granules, in a PKR-, but not RIG-I-dependent manner. Whilst stress granules were previously postulated to be essential platforms for immune activation, we found that they are not required for pro-inflammatory mediators production upon YFV infection. Collectively, our work uncovered molecular events triggered by the replication of YFV, which could prove instrumental in clarifying the pathogenesis of the disease, with possible repercussions on disease management.