Formation of cytoplasmic stress granules (SGs) and innate immune response are two distinct cellular responses to stresses. Our study investigated involvement of four innate immune proteins, retinoic acid-inducible gene I (RIG-I), melanoma differentiation-associated gene 5 (MDA5), IFN-β promoter stimulator (IPS-1) and protein kinase regulated by dsRNA (PKR) in the formation of SGs. Knockdown of IPS-1 or PKR significantly decreased the SG formation induced by dsRNA. IPS-1 depletion markedly attenuated the phosphorylation of PKR and eIF2α triggered by dsRNA, and IPS-1 facilitated the in vitro autophosphorylation of PKR. In IPS-1 depleted cells, the dsRNA-mediated association of PKR with its dsRNA binding domains or full length PKR was significantly abrogated, suggesting IPS-1 might be involved in PKR dimerization. By co-immunoprecipitation and pulldown assays, our data demonstrated that IPS-1 directly binds to PKR via its CARD domain, suggesting that effect of IPS-1 on SG formation might exert through interacting with PKR and mediating its activation. PKR was recruited into SGs upon activation while majority of IPS-1 protein formed clusters on mitochondrial membrane. Our work provides first evidence that innate signaling molecule IPS-1 plays an essential role in SG formation.

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