Glycine-Alanine Dipeptide Repeat Protein from C9-ALS Interacts with Sulfide Quinone Oxidoreductase (SQOR) to Induce the Activity of the NLRP3 Inflammasome in HMC3 Microglia: Irisflorentin Reverses This Interaction

Abstract

Amyotrophic lateral sclerosis (ALS) is a fatal rare disease of progressive degeneration of motor neurons. The most common genetic mutation in ALS is the hexanucleotide repeat expansion (HRE) located in the first intron of the C9orf72 gene (C9-ALS). HRE can produce dipeptide repeat proteins (DPRs) such as poly glycine-alanine (GA) in a repeat-associated non-ATG (RAN) translation. GA-DPR has been shown to be toxic to motor neurons in various biological models. However, its effects on microglia involved in C9-ALS have not been reported. Here, we show that GA-DPR (GA50) activates the NLR family pyrin domain containing 3 (NLRP3) inflammasome in a human HMC3 microglia model. MCC950 (specific inhibitor of the NLRP3) treatment can abrogate this activity. Next, using yeast two-hybrid screening, we identified sulfide quinone oxidoreductase (SQOR) as a GA50 interacting protein. SQOR knockdown in HMC3 cells can significantly induce the activity of the NLRP3 inflammasome by upregulating the level of intracellular reactive oxygen species and the cytoplasmic escape of mitochondrial DNA. Furthermore, we obtained irisflorentin as an effective blocker of the interaction between SQOR and GA50, thus inhibiting NLRP3 inflammasome activity in GA50-expressing HMC3 cells. These results imply the association of GA-DPR, SQOR, and NLRP3 inflammasomes in microglia and establish a treatment strategy for C9-ALS with irisflorentin.

Elevated NLRP3 Inflammasome Activation Is Associated with Motor Neuron Degeneration in ALS

“…For example, components of the NLRP3 inflammasome complex were found to be upregulated in the skeletal muscle [ 60 ], spinal cord [ 59 ], and brain [ 62 , 63 ] of SOD1 animal models. Moreover, NLRP3 inflammasome activation was detected in in vitro ALS models of TDP-43 [ 64 ] and C9Orf72 [ 65 , 66 , 67 ]. However, until now, NLRP3 inflammasome activation has not been investigated in wobbler mice, the only naturally occurring sALS animal model.…”

Mitochondria Dysfunction and Neuroinflammation in Neurodegeneration: Who Comes First?

“…The upregulation of NLRP3 inflammasome was widely demonstrated in the microglia of C9orf72- [176], hSOD1-, and hTDP-43-related ALS mouse models [177][178][179], and increased amounts of NLRP3, ASC, and caspase 1 were found both in postmortem tissue and in the serum of ALS patients [180,181]. Transcriptome analysis performed in a C9orf72 ALS mouse model and C9-ALS patients demonstrated the activation of interferon-responsive proinflammatory microglial [182].…”