Mitochondrial Inhibition by Sodium Azide Induces Assembly of eIF2α Phosphorylation-Independent Stress Granules in Mammalian Cells.

Mitochondrial stress is involved in many pathological conditions and triggers the integrated stress response (ISR). The ISR is initiated by phosphorylation of the eukaryotic translation initiation factor (eIF) 2α and results in global inhibition of protein synthesis, while the production of specific proteins important for the stress response and recovery is favored. The stalled translation preinitiation complexes phase-separate together with local RNA binding proteins into cytoplasmic stress granules (SG), which are important for regulation of cell signaling and survival under stress conditions. Here we found that mitochondrial inhibition by sodium azide (NaN3) in mammalian cells leads to translational inhibition and formation of SGs, as previously shown in yeast. Although mammalian NaN3-induced SGs are very small, they still contain the canonical SG proteins Caprin 1, eIF4A, eIF4E, eIF4G and eIF3B. Similar to FCCP and oligomycine, other mitochodrial stressors that cause SG formation, NaN3-induced SGs are formed by an eIF2α phosphorylation-independent mechanisms. Finally, we discovered that as shown for arsenite (ASN), but unlike FCCP or heatshock stress, Thioredoxin 1 (Trx1) is required for formation of NaN3-induced SGs.

Image-Based Screening for Stress Granule Regulators.

Stress granule (SG)-based RNA interference (RNAi) screening is a powerful method to discover factors that control protein synthesis and aggregation, as well as regulators of SG assembly and disassembly. Here, we describe how to set up and optimize a large-scale siRNA screen, and give a detailed outline for the automated quantification of SGs as a visual readout. Hit evaluation via calculated Z scores provides a list of candidates for further in-depth studies.

Thioredoxin 1 is required for stress granule assembly upon arsenite-induced oxidative stress.

Arsenic is a major water pollutant and health hazard, leading to acute intoxication and, upon chronic exposure, several diseases including cancer development. Arsenic exerts its pronounced cellular toxicity through its trivalent oxide arsenite (ASN), which directly inhibits numerous proteins including Thioredoxin 1 (Trx1), and causes severe oxidative stress. Cells respond to arsenic by inhibition of protein synthesis and subsequent assembly of stress granules (SGs), cytoplasmic condensates of stalled mRNAs, translation factors and RNA-binding proteins. The biological role of SGs is diverse and not completely understood; they are important for regulation of cell signaling and survival under stress conditions, and for adapting de novo protein synthesis to the protein folding capacity during the recovery from stress. In this study, we identified Trx1 as a novel component of SGs. Trx1 is required for the assembly of ASN-induced SGs, but not for SGs induced by energy deprivation or heat shock. Importantly, our results show that Trx1 is essential for cell survival upon acute exposure to ASN, through a mechanism that is independent of translation inhibition.

Dance with the Devil: Stress Granules and Signaling in Antiviral Responses.

Cells have evolved highly specialized sentinels that detect viral infection and elicit an antiviral response. Among these, the stress-sensing protein kinase R, which is activated by double-stranded RNA, mediates suppression of the host translation machinery as a strategy to limit viral replication. Non-translating mRNAs rapidly condensate by phase separation into cytosolic stress granules, together with numerous RNA-binding proteins and components of signal transduction pathways. Growing evidence suggests that the integrated stress response, and stress granules in particular, contribute to antiviral defense. This review summarizes the current understanding of how stress and innate immune signaling act in concert to mount an effective response against virus infection, with a particular focus on the potential role of stress granules in the coordination of antiviral signaling cascades.