Preventing Runaway Inflammation

Science Signaling  28 Jun 2011:
Vol. 4, Issue 179, pp. ec180
DOI: 10.1126/scisignal.4179ec180

The first response to pathogen detection involves recognition of pathogen-associated molecular patterns (PAMPs) by the Toll-like receptor (TLR) family, the RIG-1–like receptor (RLR) family, and the nucleotide-binding domain (NBD)- and leucine-rich-repeating (LRR)–containing proteins (NLRs). NLRX1 is a member of the NLR family, but in contrast to other members that promote an inflammatory response, this protein attenuates the inflammatory response to PAMPs detected by RIG-1 (see Parvatiyar and Cheng). Allen et al. and Xia et al. explored the functions of NLRX1 and its mechanisms of action and extended this protein’s role to include attenuation of TLR4 signaling, as well as RIG-1 signaling. Allen et al. generated NLRX1-knockout mice, and specific classes of cells from these animals exhibited an increased response to RIG-1 activation when treated with synthetic agonists or infected with viruses recognized by RIG-1. In the knockout cells, there was a constitutive association between RIG-1 and its adaptor MAVS, whereas in wild-type cells this interaction occurred after viral infection. Macrophages from the knockout animals did not exhibit differences in responsiveness to signals that mediate RIG-1 signaling in other cells, but these cells did show significantly increased production of interleukin-6 (IL-6) in response to activation of TLR4 by lipopolysaccharide (LPS). TRAF6, an adaptor and ubiquitin ligase involved in mediating the TLR4 signal, and the related protein TRAF3 interacted with NLRX1 in cells stimulated with LPS. Furthermore, overexpression of NLRX1 dose-dependently inhibited expression of a nuclear factor κB (NF-κB) reporter, and activation of NF-κB was enhanced in NLRX1-knockout cells.

Xia et al. identified NLRX1 in a screen for proteins that inhibited NF-κB signaling and explored the mechanistic details of this inhibition. Like Allen et al., Xia et al. reported an interaction between NLRX1 and TRAF3 or TRAF6 and found that NLRX1 also coimmunoprecipitated with the inhibitor of κB kinase (IKK) complex after LPS stimulation. In cells expressing tagged proteins, LPS stimulation resulted in K63-linked polyubiquitylation of NLRX1, which allowed the interaction of ubiquitylated NLRX1 with the IKK component NEMO, which has a ubiquitin-binding domain for K63-linked ubiquitin. Domain mapping and experiments with mutant forms of the kinases IKKα and IKKβ demonstrated that the LRR domain of NLRX1 interacted with the activated form of IKKβ, an interaction that inhibited IKKβ activity and thus phosphorylation of the IKK complex, thereby inhibiting its activity and preventing NF-κB activation. Knockdown of NLRX1 produced the expected increase in NF-κB activation in response to LPS stimulation.

Both groups examined the in vivo effects of reduced (Xia et al. with knockdown constructs) or genetic deficiency of (Allen et al.) NLRX1. Xia et al. found that the mice exhibited an increased susceptibility to death due to LPS-induced septic shock that was associated with increased circulating IL-6. Allen et al. examined the response to viral infection, including influenza viruses, and reported that although the knockout mice exhibited more rapid clearance of the virus, they had decreased survival and increased lung damage indicative of an excessive inflammatory response.

I. C. Allen, C. B. Moore, M. Schneider, Y. Lei, B. K. Davis, M. A. Scull, D. Gris, K. E. Roney, A. G. Zimmermann, J. B. Bowzard, P. Ranjan, K. M. Monroe, R. J. Pickles, S. Sambhara, J. P. Y. Ting, NLRX1 protein attenuates inflammatory responses to infection by interfering with the RIG-1-MAVS and TRAF6-NF-κB signaling pathways. Immunity 34, 854–865 (2011). [PubMed]

X. Xia, J. Cui, H. Y. Wang, L. Zhu, S. Matsueda, Q. Wang, X. Yang, J. Hong, Z. Songyang, Z. J. Chen, R.-F. Wang, NLRX1 negatively regulates TLR-induced NF-κB signaling by targeting TRAF6 and IKK. Immunity 34, 843–853 (2011). [PubMed]

K. Parvatiyar, G. Cheng, NOD so fast: NLRX1 puts the brake on inflammation. Immunity 34, 821–822 (2011). [PubMed]