Research Article

The heme-regulated inhibitor is a cytosolic sensor of protein misfolding that controls innate immune signaling

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Science  05 Jul 2019:
Vol. 365, Issue 6448, eaaw4144
DOI: 10.1126/science.aaw4144

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Linking protein misfolding and innate immunity

Multiple innate immune sensors undergo rapid assembly into large complexes known as signalosomes. This is an essential step during cellular responses to microbes and danger signals. How this process is regulated to avoid accumulation of potentially toxic protein aggregates remains poorly understood. Abdel-Nour et al. identified a pathway, dependent on heme-regulated inhibitor, eukaryotic initiation factor 2α, activating transcription factor 4, and heat shock protein B8, which controls the folding and scaffolding of innate immune sensors, allowing optimal proinflammatory signaling (see the Perspective by Pierre). The pathway appears to mirror the endoplasmic reticulum unfolded protein response (UPR), and so was named the cytosolic UPR (cUPR). The cUPR may represent a general mechanism to control protein misfolding in cells.

Science, this issue p. eaaw4144; see also p. 28

Structured Abstract

INTRODUCTION

Innate immunity relies on several families of pattern recognition molecules (PRMs) that broadly recognize microbial motifs and danger signals. Upon activation, multiple PRMs assemble into very large protein complexes or “signalosomes,” and some PRM adaptors, such as RIP2, MAVS, TRIF, and ASC, have even been shown to form amyloid-like filaments. The assembly of very large molecular structures needs to be tightly regulated to avoid accumulation of potential toxic protein aggregates, such as those causing neurodegeneration. However, the means by which PRM signalosome assembly is controlled remains poorly understood.

RATIONALE

The integrated stress response (ISR) is a highly conserved pathway that triggers eIF2α phosphorylation, a key checkpoint in the control of cellular responses to various stresses. While studying the impact of the ISR on innate immunity, we discovered that heme-regulated inhibitor (HRI), one of the four known eIF2α kinases, was essential for pro-inflammatory cytokine responses to intracellular bacterial pathogens. We sought to determine the underlying mechanism.

RESULTS

We found that HRI was critical for signaling downstream of NOD1 and NOD2, two intracellular PRMs of the nucleotide-binding domain leucine-rich repeat (NLR) protein superfamily that detect bacterial peptidoglycan and induce pro-inflammatory NF-κB signaling. Because HRI function had been previously associated with proteotoxicity, we speculated that it might control the assembly of NOD signalosomes. We indeed observed that HRI, together with the heat shock protein HSPB8, was necessary for the folding and release from endomembranes of NOD signalosomes after peptidoglycan stimulation. Presynthesized HSPB8 was released from HRI and was rapidly recruited to the PRM complex. Concomitantly, HRI was activated, triggering a pathway dependent on eIF2α, ATF4, and ATF3, which resulted in transcriptional up-regulation of HSPB8. The HRI/eIF2α/ATF4/HSPB8 signaling axis is thus critical for controlling the scaffolding of NOD signalosome and the sustained activation of NF-κB signaling. We further demonstrated that the HRI/eIF2α signaling axis was also essential for signaling downstream of MAVS and TRIF but dispensable for pathways dependent on MyD88 or STING; whether HSPB8 (or another HSP not yet identified) is involved in the HRI-dependent control of these multiple PRM pathways remains to be characterized. We further noticed that the PRM pathways regulated by HRI share the property that their adaptor proteins can form amyloid-like filaments in vitro. Indeed, overexpression of these proteins activated HRI, which suggests that potentially toxic molecular superstructures, such as self-assembling amyloid-like filaments, may be direct activators of the HRI signaling axis. In agreement with these findings, expression of α-synuclein, a protein that forms toxic amyloid filaments that are a pathological hallmark of Parkinson’s disease, induced ATF3 and HSPB8 expression through HRI.

CONCLUSION

The HRI/eIF2α/HSPB8 signaling axis identified here shares a remarkable homology with the unfolded protein response (UPR), which regulates protein folding in the endoplasmic reticulum. We propose that HRI, eIF2α, and HSPB8 define a novel cytosolic UPR (cUPR) essential for optimal innate immune signaling by large molecular platforms. Future studies should be aimed at delineating the role of the cUPR in the control and clearance of pro-aggregative proteins, such as those implicated in neurodegeneration.

The cytoplasmic unfolded protein response controls NOD1 signaling.

Upon detection of peptidoglycan (PGN), NOD1 and RIP2 undergo conformational changes, leading to the displacement of HSPB8 from HRI toward NOD1 signalosomes and subsequent NF-κB activation. Simultaneously, the dissociation of HSPB8 from HRI leads to the phosphorylation of eIF2α and the synthesis of ATF4 and ATF3, which induces de novo HSPB8 expression to sustain signaling.

Abstract

Multiple cytosolic innate sensors form large signalosomes after activation, but this assembly needs to be tightly regulated to avoid accumulation of misfolded aggregates. We found that the eIF2α kinase heme-regulated inhibitor (HRI) controls NOD1 signalosome folding and activation through a process requiring eukaryotic initiation factor 2α (eIF2α), the transcription factor ATF4, and the heat shock protein HSPB8. The HRI/eIF2α signaling axis was also essential for signaling downstream of the innate immune mediators NOD2, MAVS, and TRIF but dispensable for pathways dependent on MyD88 or STING. Moreover, filament-forming α-synuclein activated HRI-dependent responses, which suggests that the HRI pathway may restrict toxic oligomer formation. We propose that HRI, eIF2α, and HSPB8 define a novel cytosolic unfolded protein response (cUPR) essential for optimal innate immune signaling by large molecular platforms, functionally homologous to the PERK/eIF2α/HSPA5 axis of the endoplasmic reticulum UPR.

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