Toll-like receptors promote mitochondrial translocation of nuclear transcription factor nuclear factor of activated T-cells in prolonged microglial activation

Microglia are resident macrophages in the CNS that scavenge pathogens, dying cells, and molecules using pattern recognition Toll-like receptors (TLRs). Nuclear factor of activated T-cells (NFAT) family transcription factors also regulate inflammatory responses in microglia. However, whether there exists cross talk between TLR and NFAT signaling is unclear. Here we show that chronic activation of murine microglia by prolonged stimulation of Toll-like receptor 4 (TLR4) ligand lipopolysaccharides (LPSs) leads to unexpected translocation of NFAT1 into mitochondria. This mitochondrial import of NFAT1 is independent of calcium/calcineurin signaling. Instead, inhibition of Toll/interleukin 1 receptor domain-containing adapter-inducing interferon-β (TRIF) pathway blocks the mitochondrial translocation of NFAT1. Functionally, inhibition of NFAT1 reduces the TRIF-mediated expression of interferon-β and compromises the production of ATP and reactive oxygen species in LPS-treated microglia. Therefore, our findings reveal a new inflammatory signaling pathway that links TLR with NFAT in regulating cytokine production and mitochondrial activity during chronic microglial activation. Significance Statement: Nuclear factor of activated T-cells (NFAT) family transcription factors are known to undergo nuclear translocation in response to inflammatory stimulation. In this study, we uncovered a surprise transportation of NFATs into mitochondria in microglia after a prolonged treatment with bacteria endotoxin lipopolysaccharides (LPSs). LPSs activated Toll-like receptor 4 and its downstream Toll/interleukin 1 receptor-domain-containing adapter-inducing interferon-β (TRIF) to regulate the mitochondrial translocation of NFAT in microglia, whereas genetic inhibition of NFAT1 compromised TRIF-mediated cytokine production and reduced ATP and reactive oxygen species generation. These findings reveal a previously undescribed mitochondrial translocation of NFAT in microglia responding to extended activation of Toll-like receptor-mediated signaling transduction pathways.