Zika Virus Hijacks Stress Granule Proteins and Modulates the Host Stress Response

Abstract

Zika virus (ZIKV), a member of the Flaviviridae family, has recently emerged as an important human pathogen with increasing economic and health impact worldwide. Because of its teratogenic nature and association with the serious neurological condition Guillain-Barré syndrome, a tremendous amount of effort has focused on understanding ZIKV pathogenesis. To gain further insights into ZIKV interaction with host cells, we investigated how this pathogen affects stress response pathways. While ZIKV infection induces stress signaling that leads to phosphorylation of eIF2␣ and cellular translational arrest, stress granule (SG) formation was inhibited. Further analysis revealed that the viral proteins NS3 and NS4A are linked to translational repression, whereas expression of the capsid protein, NS3/NS2B-3, and NS4A interfered with SG formation. Some, but not all, flavivirus capsid proteins also blocked SG assembly, indicating differential interactions between flaviviruses and SG biogenesis pathways. Depletion of the SG components G3BP1, TIAR, and Caprin-1, but not TIA-1, reduced ZIKV replication. Both G3BP1 and Caprin-1 formed complexes with capsid, whereas viral genomic RNA stably interacted with G3BP1 during ZIKV infection. Taken together, these results are consistent with a scenario in which ZIKV uses multiple viral components to hijack key SG proteins to benefit viral replication.IMPORTANCE There is a pressing need to understand ZIKV pathogenesis in order to advance the development of vaccines and therapeutics. The cellular stress response constitutes one of the first lines of defense against viral infection; therefore, understanding how ZIKV evades this antiviral system will provide key insights into ZIKV biology and potentially pathogenesis. Here, we show that ZIKV induces the stress response through activation of the UPR (unfolded protein response) and PKR (protein kinase R), leading to host translational arrest, a process likely mediated by the viral proteins NS3 and NS4A. Despite the activation of translational shutoff, formation of SG is strongly inhibited by the virus. Specifically, ZIKV hijacks the core SG proteins G3BP1, TIAR, and Caprin-1 to facilitate viral replication, resulting in impaired SG assembly. This process is potentially facilitated by the interactions of the viral RNA with G3BP1 as well as the viral capsid protein with G3BP1 and Caprin-1. Interestingly, expression of capsid proteins from several other flaviviruses also inhibited SG formation. Taken together, the present study provides novel insights into how ZIKV modulates cellular stress response pathways during replication.

Endoplasmic reticulum: a focal point of Zika virus infection

“…Consistent with this finding, pharmacological inhibition of GADD34-mediated eIF2α dephosphorylation rescued SGs assembly and decreased ZIKV particles production [77]. Additionally, another study reported that ZIKV proteins, namely capsid, NS3, NS2B-NS3, and NS4A proteins, suppressed SGs assembly; however, this modulation was observed in an eIF2α-independent manner [78]. ZIKV capsid inhibited SGs assembly by forming stable complexes with SG core proteins, Ras GTPase-activating proteinbinding protein 1 (G3BP1) and caprin-1, but not with T-cell-restricted intracellular antigen 1 related (TIAR) protein [78].…”

“…Additionally, another study reported that ZIKV proteins, namely capsid, NS3, NS2B-NS3, and NS4A proteins, suppressed SGs assembly; however, this modulation was observed in an eIF2α-independent manner [78]. ZIKV capsid inhibited SGs assembly by forming stable complexes with SG core proteins, Ras GTPase-activating proteinbinding protein 1 (G3BP1) and caprin-1, but not with T-cell-restricted intracellular antigen 1 related (TIAR) protein [78]. ZIKV utilized these SG core proteins for viral protein and virion production.…”

“…Beside regulating eIF2α phosphorylation and hijacking key SG proteins, RNA viruses were reportedly capable of interfering SGs assembly through cleaving and redistributing SGs nucleating factors [74]. It was previously reported that ZIKV NS2B-NS3 protease could cleave host antiviral factors to impair intrinsic host defense; intriguingly, ZIKV did not mediate the cleavage of SG factors even though SGs assembly was affected by ZIKV NS3 and NS2B-NS3 protease [77][78][79][80][81]. Nonetheless, ZIKV infection was found to facilitate the redistribution of TIAR to viral replication sites, which correlates to viral replication output [77,78,81].…”

Zika Virus Production Is Resistant to RNase L Antiviral Activity

“…Degradation of cellular RNA by RNase L inhibits global protein translation to limit viral infection, although many viruses that induce cellular protein translation arrest have found ways to concurrently maintain viral protein synthesis (29,30). Inhibition of protein translation during ZIKV infection has been previously described (29,31); we therefore investigated whether translation arrest was mediated by RNase L, and, if so, whether ZIKV nascent protein synthesis was affected by activation of RNase L. We infected WT or RNase L KO A549 cells with ZIKV, and at different time points postinfection treated cells with puromycin, which is incorporated into nascent protein, before harvesting whole-cell lysate. Coomassie blue staining for preexisting total cellular and viral protein, as well as Western blotting for GAPDH (glyceraldehyde-3-phosphate dehydrogenase) protein, from both WT and RNase L KO cells over time confirmed equal amounts of total protein abundance in each sample ( Fig.…”

Protein Phase Separation: A New Phase in Cell Biology

“…Moreover, some viruses, such as flaviviruses, including Zika, even hijack SG proteins to aid in their replication [128,129]. Although still in its infancy, we would argue that since protein aggregation and phase separation are implicated in numerous human pathological conditions, a better understanding of these processes will help us develop novel therapeutic strategies in the wider field of human medicine.…”