Researchers have uncovered an immune mechanism by which host cells combat bacterial infection, and at the same time found that a protein crucial to that process can sense and respond to misfolded proteins in all mammalian cells.
The protein is called heme-regulated inhibitor or HRI, and the researchers showed that during bacterial infection it triggers and coordinates a chain reaction among other proteins that form a larger complex. That larger group or 'signalosome' amplifies inflammation and leads to an anti-bacterial response.
The authors found that HRI was critical for signaling downstream of NOD1 and NOD2, two intracellular pattern recognition molecules (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, authors speculated that it might control the assembly of NOD signalosomes.
They 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.
The authors 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.
But HRI can also regulate protein folding in other cell types, the researchers showed. Protein folding, which helps determine the 3-D shape of a protein and is essential for its function, is implicated in non-infectious diseases including the neurodegenerative disorders Parkinson's, Alzheimer's and ALS.
"The innate immune function that we discovered is essentially a mechanism of protein scaffolding, which is important because you want a quick and orderly response to bacterial infection," says the senior author. "But we also found that same pathway is important for protein scaffolding and aggregation in other cells, which opens promising research angles for neurodegenerative and other diseases."The journal Science published the findings.
The authors 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.
Researchers have studied HRI for over three decades, but mostly in the context of red blood cell disorders. "This protein appears in all cells in the mammalian body and was recognized as a broad or promiscuous sensor," says the lead author on the paper. "But it was overlooked relative to pattern recognition molecules and the formation of amyloid-like structures. We had to test its role in several different pathways before we believed what we saw."
The researchers developed a novel technique to study the effects of HRI. They adapted a biochemistry assay which helped them distinguish between folded and misfolded proteins when looking at protein aggregates. Scientists have struggled to make that distinction in part because most available tests only work in test tubes and are not adaptable to cells.
The researchers have early pre-clinical data that shows HRI could protect against the type of neurodegeneration seen in Parkinson's. "Speculatively, it might be possible to find molecules that produce HRI's protective effects, which could lead to a bona fide therapy," says another author. Current therapies for Parkinson's focus on finding and clearing out protein aggregates, rather than fixing cellular defects before those clusters accumulate.
The senior author says, "We are focused on Parkinson's because it's a very important disease for human health, and because its hallmark is protein aggregation inside cells, so it may be a perfect model to test this new pathway."
Next steps include biochemical investigations of HRI and related complexes during protein misfolding, and animal studies of neurodegenerative disease to further validate the new pathway, which shares many features with a similar pathway in humans.