Toll-like receptors (TLRs) are major players in the innate immune system-recognizing pathogens and differentiating self/non-self components of immunity. These proteins are present either on the plasma membrane or endosome and recognize pathogens at their extracellular domains. They are characterized by a single transmembrane helix and an intracellular toll-interleukin-1 receptor (TIR) domain. Few TIRs directly invoke downstream signaling, while others require other TIR domains of adaptors like TIR domain-containing adaptor-inducing interferon-β (TRIF) and TRIF-related adaptor molecule (TRAM). On recognizing pathogenic lipopolysaccharides, TLR4 dimerises and interacts with the intracellular TRAM dimer through the TIR domain to recruit a downstream signaling adaptor (TRIF). We have performed an in-depth study of the structural effect of two mutations (P116H and C117H) at the dimeric interface of the adaptor TRAM, which are known to abrogate downstream signaling. We modeled the structure and performed molecular dynamics studies in order to decipher the structural basis of this effect. We observed that these mutations led to an increased radius of gyration of the complex and resulted in several changes to the interaction energy values when compared against the wild type (WT) and positive control mutants. We identified highly interacting residues as hubs in the WT dimer, and a few such hubs that were lost in the mutant dimers. Changes in the protein residue path, hampering the information flow between the crucial A86/E87/D88/D89 and T155/S156 sites, were observed for the mutants. Overall, we show that such residue changes can have subtle but long-distance effects, impacting the signaling path allosterically.

Previous research results of our group showed that Toll-like receptor 4 (TLR4) and nucleolin synergistically mediate respiratory syncytial virus (RSV) infection in human central neuron cells, but the specific mechanism remains unclear. Here we designed and synthesized lentiviruses with TIR (674-815 aa), TLR4 (del 674-815 aa), GAR (645-707 aa), and NCL (del 645-707 aa) domains, and obtained stable overexpression cell lines by drug screening, and subsequently infected RSV at different time points. Laser confocal microscopy and coimmunoprecipitation were used for the observation of co-localization and interaction of TIR/GAR domains. Western blot analysis was used for the detection of p-NF-κB and LC3 protein expression. Real-time PCR was used for the detection of TLR4/NCL mRNA expression. ELISA assay was used to measure IL-6, IL-1β, and TNF-α concentrations and flow cytometric analysis was used for the study of apoptosis. Our results suggest that overexpression of TIR and GAR domains can exacerbate apoptosis and autophagy, and that TIR and GAR domains can synergistically mediate RSV infection and activate the NF-κB signaling pathway, which regulates the secretion of downstream inflammatory factors, such as IL-6, IL-1β, and TNF-α, and ultimately leads to neuronal inflammatory injury.

Plants deploy cell-surface and intracellular receptors to detect pathogen attack and trigger innate immune responses. Inside host cells, families of nucleotide-binding/leucine-rich repeat (NLR) proteins serve as pathogen sensors or downstream mediators of immune defence outputs and cell death, which prevent disease. Established genetic underpinnings of NLR-mediated immunity revealed various strategies plants adopt to combat rapidly evolving microbial pathogens. The molecular mechanisms of NLR activation and signal transmission to components controlling immunity execution were less clear. Here, we review recent protein structural and biochemical insights to plant NLR sensor and signalling functions. When put together, the data show how different NLR families, whether sensors or signal transducers, converge on nucleotide-based second messengers and cellular calcium to confer immunity. Although pathogen-activated NLRs in plants engage plant-specific machineries to promote defence, comparisons with mammalian NLR immune receptor counterparts highlight some shared working principles for NLR immunity across kingdoms.

Sepsis is a dysregulated systemic inflammatory response caused by infection that leads to multiple organ injury and high mortality without effective treatment. Corilagin, a natural polyphenol extracted from traditional Chinese herbs, exhibits strong anti-inflammatory properties. However, the role for Corilagin in lipopolysaccharide (LPS)-induced sepsis and the molecular mechanisms underlying this process have not been completely explored. Here we determine the effect of Corilagin on LPS-treated mice and use a screening approach integrating surface plasmon resonance with liquid chromatography-tandem mass spectrometry (SPR-LC-MS/MS) to further explore the therapeutic target of Corilagin. We discovered that Corilagin significantly prolonged the survival time of septic mice, attenuated the multi-organ injury and the expression of pyroptosis-related proteins in tissues of LPS-treated mice. In vitro studies revealed that Corilagin inhibited pyroptosis and NLRP3 inflammasome activation in LPS-treated macrophages followed with ATP stimulation, as reflected by decreased levels of GSDMD-NT and activated caspase-1, and reduced ASC specks formation. Mechanistically, Corilagin alleviated the formation of ASC specks and blocked the interaction of ASC and pro-caspase1 by competitively binding with the caspase recruitment domain (CARD) of ASC. Additionally, Corilagin interrupted the TLR4-MyD88 interaction through targeting TIR domain of MyD88, leading to the inhibition of NF-κB activation and NLRP3 production. In addition, Corilagin downregulated genes associated with several inflammatory responses and inflammasome-related signaling pathways in LPS-stimulated macrophages. Overall, our results indicate that the inhibitory effect of Corilagin on pyroptosis through targeting TIR domain of MyD88 and binding the CARD domain of ASC in macrophages plays an essential role in protection against LPS-induced sepsis.

While working for the United States Department of Agriculture on the North Dakota Agricultural College campus in Fargo, North Dakota, in the 1940s and 1950s, Harold H. Flor formulated the genetic principles for coevolving plant host-pathogen interactions that govern disease resistance or susceptibility. His \'gene-for-gene\' legacy runs deep in modern plant pathology and continues to inform molecular models of plant immune recognition and signaling. In this review, we discuss recent biochemical insights to plant immunity conferred by nucleotide-binding domain/leucine-rich-repeat (NLR) receptors, which are major gene-for-gene resistance determinants in nature and cultivated crops. Structural and biochemical analyses of pathogen-activated NLR oligomers (resistosomes) reveal how different NLR subtypes converge in various ways on calcium (Ca2+) signaling to promote pathogen immunity and host cell death. Especially striking is the identification of nucleotide-based signals generated enzymatically by plant toll-interleukin 1 receptor (TIR) domain NLRs. These small molecules are part of an emerging family of TIR-produced cyclic and noncyclic nucleotide signals that steer immune and cell-death responses in bacteria, mammals, and plants. A combined genetic, molecular, and biochemical understanding of plant NLR activation and signaling provides exciting new opportunities for combatting diseases in crops. [Formula: see text] Copyright © 2023 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Toll-like receptors (TLRs) are pattern recognition receptors present on the surface of cells playing a crucial role in innate immunity. One of the TLRs, TLR4, recognizes LPS (Lipopolysaccharide) as its ligand leading to the release of anti-inflammatory mediators as well as pro-inflammatory cytokines through signal transduction and domain recruitment. TLR4 homodimerizes at its intracellular TIR (Toll/interleukin-1 receptor) domain that helps in the recruitment of the TRAM/TICAM2 (TIR domain-containing adaptor molecule 2) molecule. TRAM also contains TIR domain which in turn, dimerizes and functions as an adapter protein to further recruit TRIF/TICAM1 (TIR domain-containing adaptor molecule 1) protein for mediating downstream signaling. Apart from LPS, TLR4 also recognizes endogenous ligands like fibrinogen, HMGB1, and hyaluronan in autoimmune conditions and sepsis. We employed computational approaches to target TRAM and recognize small molecule inhibitors from small molecules of natural origin, as contained in the Super Natural II database. Finally, cell reporter assays and NMR studies enabled the identification of promising lead compounds. Hence, this study aims to attenuate the signaling of the TLR4-TRAM-TRIF cascade in these auto-inflammatory conditions.

Plant disease resistance involves both detection of microbial molecular patterns by cell-surface pattern recognition receptors and detection of pathogen effectors by intracellular NLR immune receptors. NLRs are classified as sensor NLRs, involved in effector detection, or helper NLRs required for sensor NLR signaling. TIR-domain-containing sensor NLRs (TNLs) require helper NLRs NRG1 and ADR1 for resistance, and helper NLR activation of defense requires the lipase-domain proteins EDS1, SAG101, and PAD4. Previously, we found that NRG1 associates with EDS1 and SAG101 in a TNL activation-dependent manner [X. Sun et al., Nat. Commun. 12, 3335 (2021)]. We report here how the helper NLR NRG1 associates with itself and with EDS1 and SAG101 during TNL-initiated immunity. Full immunity requires coactivation and mutual potentiation of cell-surface and intracellular immune receptor-initiated signaling [B. P. M. Ngou, H.-K. Ahn, P. Ding, J. D. G. Jones, Nature 592, 110-115 (2021), M. Yuan et al., Nature 592, 105-109 (2021)]. We find that while activation of TNLs is sufficient to promote NRG1-EDS1-SAG101 interaction, the formation of an oligomeric NRG1-EDS1-SAG101 resistosome requires the additional coactivation of cell-surface receptor-initiated defense. These data suggest that NRG1-EDS1-SAG101 resistosome formation in vivo is part of the mechanism that links intracellular and cell-surface receptor signaling pathways.

In 2011, the Nobel Prize in Physiology or Medicine was awarded to three immunologists: Bruce A. Beutler, Jules A. Hoffmann, and Ralph M. Steinman. While Steinman was honored for his work on dendritic cells and adaptive immunity, Beutler and Hoffman received the prize for their contributions to discoveries in innate immunity. In 1996, Hoffmann found the toll gene to be crucial for mounting antimicrobial responses in fruit flies, first implicating this developmental gene in immune signaling. Two years later, Beutler built on this observation by describing a Toll-like gene, tlr4, as the receptor for the bacterial product LPS, representing a crucial step in innate immune activation and protection from bacterial infections in mammals. These publications spearheaded research in innate immune sensing and sparked a huge interest regarding innate defense mechanisms in the following years and decades. Today, Beutler and Hoffmann\'s research has not only resulted in the discovery of the role of multiple TLRs in innate immunity but also in a much broader understanding of the molecular components of the innate immune system. In this review, we aim to collect the discoveries leading up to the publications of Beutler and Hoffmann, taking a close look at how early advances in both developmental biology and immunology converged into the research awarded with the Nobel Prize. We will also discuss how these discoveries influenced future research and highlight the importance they hold today.

Axons are considered to be particularly vulnerable components of the nervous system; impairments to a neuron\'s axon leads to an effective silencing of a neuron\'s ability to communicate with other cells. Nervous systems have therefore evolved plasticity mechanisms for adapting to axonal damage. These include acute mechanisms that promote the degeneration and clearance of damaged axons and, in some cases, the initiation of new axonal growth and synapse formation to rebuild lost connections. Here we review how these diverse processes are influenced by the therapeutically targetable enzyme SARM1. SARM1 catalyzes the breakdown of NAD+, which, when unmitigated, can lead to rundown of this essential metabolite and axonal degeneration. SARM1\'s enzymatic activity also triggers the activation of downstream signaling pathways, which manifest numerous functions for SARM1 in development, innate immunity and responses to injury. Here we will consider the multiple intersections between SARM1 and the injury signaling pathways that coordinate cellular adaptations to nervous system damage.

The NADase SARM1 (sterile alpha and TIR motif containing 1) is a key executioner of axon degeneration and a therapeutic target for several neurodegenerative conditions. We show that a potent SARM1 inhibitor undergoes base exchange with the nicotinamide moiety of nicotinamide adenine dinucleotide (NAD+) to produce the bona fide inhibitor 1AD. We report structures of SARM1 in complex with 1AD, NAD+ mimetics and the allosteric activator nicotinamide mononucleotide (NMN). NMN binding triggers reorientation of the armadillo repeat (ARM) domains, which disrupts ARM:TIR interactions and leads to formation of a two-stranded TIR domain assembly. The active site spans two molecules in these assemblies, explaining the requirement of TIR domain self-association for NADase activity and axon degeneration. Our results reveal the mechanisms of SARM1 activation and substrate binding, providing rational avenues for the design of new therapeutics targeting SARM1.