Title

NLRX1 modulates differentially NLRP3 inflammasome activation and NF-kappaB signaling during Fusobacterium nucleatum infection

Introduction/Context

NOD-like receptors (NLRs) play a significant role in regulation of host innate immunity, yet their role in periodontitis remains to be defined. NLRX1, a member of the NLR family that localizes to mitochondria, enhances mitochondrial ROS (mROS) generation. mROS can activate the NLRP3 inflammasome, yet the role of NLRX1 in NLRP3 inflammasome activation has not been examined. In this study, we revealed the mechanism by which NLRX1 positively regulates ATP-induced NLRP3 inflammasome activation through mROS in gingival epithelial cells (GECs).

Method

GECs were plated at 80% confluency and infected with F. nucleatum at an MOI of 100 for 24h and treated with 5mM ATP during the last 1h of infection where indicated. NLRX1 and control knock-downs in GECS were generated using lentivirus infection and selection of sh_RNA-expressing cells. NF-κB activation, mROS generation, ASC specks formation were detected and quantified by fluorescence microscopy. Caspase-1 activation was detected by western blotting, IL-8 production was measured using Real-time PCR.

Results

We found that depletion of NLRX1 by shRNA attenuated ATP-induced mROS generation and redistribution of the NLRP3 inflammasome adaptor protein, ASC. Furthermore, depletion of NLRX1 inhibited Fusobacterium nucleatum infection-activated caspase-1, suggesting that it also inhibits the NLRP3 inflammasome. Conversely, NLRX1 also acted as a negative regulator of NF-κB signaling and IL-8 expression.

Significance/Conclusions

NLRX1 stimulates detection of the pathogen F. nucleatum via the inflammasome, while dampening cytokine production. We expect that commensals should not activate the inflammasome, and NLRX1 should decrease their ability to stimulate expression of pro-inflammatory cytokines such as IL-8. Therefore, NLRX1 may act as a potential switch with regards to anti-microbial responses in healthy or diseased states in the oral cavity.

Comments

This work was supported by the National Institutes of Health, FAPERJ scholarship to CLCAS, and Funds from the Arthur Dugoni School of Dentistry.

Location

University of the Pacific, Dugoni Dental School, San Francisco, CA

Format

Poster

Poster Session

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NLRX1 modulates differentially NLRP3 inflammasome activation and NF-kappaB signaling during Fusobacterium nucleatum infection

University of the Pacific, Dugoni Dental School, San Francisco, CA

NOD-like receptors (NLRs) play a significant role in regulation of host innate immunity, yet their role in periodontitis remains to be defined. NLRX1, a member of the NLR family that localizes to mitochondria, enhances mitochondrial ROS (mROS) generation. mROS can activate the NLRP3 inflammasome, yet the role of NLRX1 in NLRP3 inflammasome activation has not been examined. In this study, we revealed the mechanism by which NLRX1 positively regulates ATP-induced NLRP3 inflammasome activation through mROS in gingival epithelial cells (GECs).