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W. Gu

Bio: W. Gu is an academic researcher from University of Queensland. The author has contributed to research in topics: NAD+ kinase & Medicine. The author has an hindex of 3, co-authored 8 publications receiving 222 citations.

Papers
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Journal ArticleDOI
23 Aug 2019-Science
TL;DR: NAD depletion as pathogen response One way that plants respond to pathogen infection is by sacrificing the infected cells, and Toll/interleukin-1 receptor (TIR) domains cleave the metabolic cofactor nicotinamide adenine dinucleotide (NAD+) as part of their cell-death signaling in response to pathogens.
Abstract: SARM1 (sterile alpha and TIR motif containing 1) is responsible for depletion of nicotinamide adenine dinucleotide in its oxidized form (NAD+) during Wallerian degeneration associated with neuropathies. Plant nucleotide-binding leucine-rich repeat (NLR) immune receptors recognize pathogen effector proteins and trigger localized cell death to restrict pathogen infection. Both processes depend on closely related Toll/interleukin-1 receptor (TIR) domains in these proteins, which, as we show, feature self-association-dependent NAD+ cleavage activity associated with cell death signaling. We further show that SARM1 SAM (sterile alpha motif) domains form an octamer essential for axon degeneration that contributes to TIR domain enzymatic activity. The crystal structures of ribose and NADP+ (the oxidized form of nicotinamide adenine dinucleotide phosphate) complexes of SARM1 and plant NLR RUN1 TIR domains, respectively, reveal a conserved substrate binding site. NAD+ cleavage by TIR domains is therefore a conserved feature of animal and plant cell death signaling pathways.

326 citations

Journal ArticleDOI
07 Apr 2021-Neuron
TL;DR: In this paper, the authors demonstrate that SARM1 is activated by an increase in the ratio of nicotinamide mononucleotide (NMN) to NAD+ and show that both metabolites compete for binding to the auto-inhibitory N-terminal armadillo repeat (ARM) domain.

141 citations

Journal ArticleDOI
TL;DR: In this article , the orthosteric site of SARM1 spans two TIR domain molecules, and in the activated state, TIR domains self-associate to form two-stranded assemblies.

43 citations

Journal ArticleDOI
01 Sep 2022-Science
TL;DR: 3'cADPR is an activator of ThsA effector proteins from bacterial anti-phage defense systems termed Thoeris, and a suppressor of plant immunity when produced by the effector HopAM1.
Abstract: Cyclic adenosine diphosphate (ADP)–ribose (cADPR) isomers are signaling molecules produced by bacterial and plant Toll/interleukin-1 receptor (TIR) domains via nicotinamide adenine dinucleotide (oxidized form) (NAD+) hydrolysis. We show that v-cADPR (2′cADPR) and v2-cADPR (3′cADPR) isomers are cyclized by O-glycosidic bond formation between the ribose moieties in ADPR. Structures of 2′cADPR-producing TIR domains reveal conformational changes that lead to an active assembly that resembles those of Toll-like receptor adaptor TIR domains. Mutagenesis reveals a conserved tryptophan that is essential for cyclization. We show that 3′cADPR is an activator of ThsA effector proteins from the bacterial antiphage defense system termed Thoeris and a suppressor of plant immunity when produced by the effector HopAM1. Collectively, our results reveal the molecular basis of cADPR isomer production and establish 3′cADPR in bacteria as an antiviral and plant immunity–suppressing signaling molecule. Description Production of small signaling isomers Isomers of the small signaling molecule cyclic ADP ribose (cADPR) are produced when bacteria and plants respond to pathogens. Manik et al. found that cyclization is managed by Toll/interleukin-1 receptor (TIR) domain proteins that are themselves activated by conformational changes. One cyclic isomer, 3′cADPR, activates bacterial anti-hage defense systems in bacteria and suppresses immunity in plants. —PJH The production of signals in bacterial and plant immune responses reveals diversity in small signaling molecules. INTRODUCTION Organisms from bacteria to animals and plants must defend themselves against pathogens. Homologous protein motifs exist in immune pathways of all organisms. One such motif is the TIR domain, named after the mammalian immune receptors—Toll-like receptors and interleukin-1 receptors—where it was first identified. Two properties are shared among most TIR domains from all organisms: the ability to self-associate and enzymatic activity involving the cleavage of nicotinamide adenine dinucleotide (oxidized form) (NAD+). NAD+ is a metabolite with redox properties that has roles in many cellular processes. In some cases, cleavage of NAD+ leads to the production of cyclic adenosine monophosphate (ADP)–ribose (cADPR) isomers. RATIONALE In bacteria, NAD+-cleavage activity by TIR domain–containing proteins plays a role in defense signaling, as well as suppression of host immunity. One corresponding pathway is termed the Thoeris defense system. This signaling pathway protects bacteria against phage infection and involves the thsA and thsB genes. Upon phage infection, ThsB (a TIR-domain protein) cleaves NAD+ and produces a cADPR isomer, which activates ThsA-mediated killing of the infected cell, thus protecting the bacterial population. Another bacterial protein that produces a cADPR isomer is HopAM1, the TIR-domain effector protein from Pseudomonas syringae DC3000, which is involved in suppressing plant immunity. The chemical structures and mechanisms of action of the responsible cADPR isomers were unknown before this work. Our aim was to determine the chemical structures of cADPR isomers, the structural basis of their production by bacterial TIR domains, and their mechanism of action in Thoeris defense signaling and suppression of plant immunity. RESULTS Using a combination of methods, including nuclear magnetic resonance (NMR), mass spectrometry, and crystallography, we show that the cADPR isomers are cyclized by O-glycosidic bond formation between the ribose moieties in ADPR. Structures of TIR domains that produce cADPR isomers, as determined by crystallography and cryo–electron microscopy, reveal conformational changes that lead to an active assembly that resembles those of Toll-like receptor adaptor TIR domains. Mutagenesis reveals a conserved tryptophan that is essential for cyclization. Using crystallography and biophysical approaches, we show that one of the cADPR isomers (3′cADPR) is an activator of Thoeris ThsA proteins responsible for antiphage defense, by inducing a change in its tetrameric state. We also show that the same cADPR isomer is a suppressor of plant immunity when produced by the effector HopAM1. CONCLUSION Collectively, our results reveal the molecular basis of cADPR isomer production. The 2′cADPR and 3′cADPR differ only in the location of the O-glycosidic bond between the ribose moieties in ADPR. These compounds add to the growing list of signaling molecules identified in immune pathways that involve proteins containing TIR domains and may represent intermediates in their synthesis or signaling molecules with their own distinctive activities. Our results establish the 3′cADPR isomer produced by bacterial TIR domain–containing proteins as an antiviral and plant immunity–suppressing signaling molecule. Diverse immune roles of bacterial cADPR isomers. Bacteria have TIR domain–containing proteins that cleave NAD to produce cyclic ADPR isomers with different cyclic linkages. One of these molecules, 3′cADPR, has roles in diverse immunity pathways. It acts as an activator of the Thoeris antiphage defense system by binding to the protein ThsA. When produced by the effector HopAM1 from the plant pathogen P. syringae, it suppresses plant immunity.

29 citations

Journal ArticleDOI
TL;DR: The axon survival factor NMNAT2 and pro-degenerative factor SARM1 have been extensively characterized and plays an essential role in maintaining the axon integrity as mentioned in this paper, which can be activated in necroptosis and in genetic, toxic or metabolic disorders, physical injury and neuroinflammation.
Abstract: Axon degeneration represents a pathological feature of many neurodegenerative diseases, including Alzheimer’s disease and Parkinson’s disease where axons die before the neuronal soma, and axonopathies, such as Charcot-Marie-Tooth disease and hereditary spastic paraplegia. Over the last two decades, it has slowly emerged that a central signaling pathway forms the basis of this process in many circumstances. This is an axonal NAD-related signaling mechanism mainly regulated by the two key proteins with opposing roles: the NAD-synthesizing enzyme NMNAT2, and SARM1, a protein with NADase and related activities. The crosstalk between the axon survival factor NMNAT2 and pro-degenerative factor SARM1 has been extensively characterized and plays an essential role in maintaining the axon integrity. This pathway can be activated in necroptosis and in genetic, toxic or metabolic disorders, physical injury and neuroinflammation, all leading to axon pathology. SARM1 is also known to be involved in regulating innate immunity, potentially linking axon degeneration to the response to pathogens and intercellular signaling. Understanding this NAD-related signaling mechanism enhances our understanding of the process of axon degeneration and enables a path to the development of drugs for a wide range of neurodegenerative diseases.

18 citations


Cited by
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Journal ArticleDOI
28 May 2020-Cell
TL;DR: Recent advances in understanding the mechanisms underlying activation of the main classes of immune receptors are highlighted, the current understanding of their signaling mechanisms are summarized, and an updated model for SA perception and signaling is discussed.

399 citations

Posted ContentDOI
10 Apr 2020-bioRxiv
TL;DR: This study supports an alternative model in which PTI is in fact an indispensable component of ETI during bacterial infection, implying that ETI halts pathogen infection, in part, by directly co-opting the anti-pathogen mechanisms proposed for PTI.
Abstract: The plant immune system is fundamental to plant survival in natural ecosystems and productivity in crop fields. Substantial evidence supports the prevailing notion that plants possess a two-tiered innate immune system, called pattern-triggered immunity (PTI) and effector-triggered immunity (ETI). PTI is triggered by microbial patterns via cell surface-localized pattern-recognition receptors (PRRs), whereas ETI is activated by pathogen effector proteins via mostly intracellularly-localized receptors called nucleotide-binding, leucine-rich repeat proteins (NLRs)1-4. PTI and ETI are initiated by dist 30 inct activation mechanisms and are considered to act independently and have evolved sequentially5,6. Here we show that, contrary to the perception of PTI and ETI being separate immune signaling pathways, Arabidopsis PRR/co-receptor mutants, fls2/efr/cerk1 and bak1/bkk1/cerk1 triple mutants, are greatly impaired in ETI responses when challenged with incompatible Pseudomonas syrinage bacteria. We further show that the NADPH oxidase (RBOHD)-mediated production of reactive oxygen species (ROS) is a critical early signaling event connecting PRR and NLR cascades and that PRR-mediated phosphorylation of RBOHD is necessary for full activation of RBOHD during ETI. Furthermore, NLR signaling rapidly augments the transcript and protein levels of key PTI components at an early stage and in a salicylic acid-independent manner. Our study supports an alternative model in which PTI is in fact an indispensable component of ETI during bacterial infection, implying that ETI halts pathogen infection, in part, by directly co-opting the anti-pathogen mechanisms proposed for PTI. This alternative model conceptually unites two major immune signaling pathways in the plant kingdom and mechanistically explains the long-observed similarities in downstream defense outputs between PTI and ETI.

274 citations

Journal ArticleDOI
04 Dec 2020-Science
TL;DR: The mechanism of pathogen effector recognition by plant innate immune receptors and the formation of a signaling-active complex are elucidated and a deeper understanding of the principles that govern nonself recognition by NLRs and their activation of innate immune responses is explained.
Abstract: Direct or indirect recognition of pathogen-derived effectors by plant nucleotide-binding leucine-rich repeat (LRR) receptors (NLRs) initiates innate immune responses. The Hyaloperonospora arabidopsidis effector ATR1 activates the N-terminal Toll-interleukin-1 receptor (TIR) domain of Arabidopsis NLR RPP1. We report a cryo-electron microscopy structure of RPP1 bound by ATR1. The structure reveals a C-terminal jelly roll/Ig-like domain (C-JID) for specific ATR1 recognition. Biochemical and functional analyses show that ATR1 binds to the C-JID and the LRRs to induce an RPP1 tetrameric assembly required for nicotinamide adenine dinucleotide hydrolase (NADase) activity. RPP1 tetramerization creates two potential active sites, each formed by an asymmetric TIR homodimer. Our data define the mechanism of direct effector recognition by a plant NLR leading to formation of a signaling-active holoenzyme.

244 citations

Journal ArticleDOI
TL;DR: In this article, the relationship between the two layers of plant innate immunity is discussed, and the existence of intricate interactions between PRR-mediated and intracellular nucleotide-binding domain leucine-rich repeat containing receptors (NLRs) are investigated.

241 citations

Journal ArticleDOI
04 Dec 2020-Science
TL;DR: The structure of the ROQ1 (recognition of XopQ 1)–XopQ (Xanthomonas outer protein Q) complex was used as a model to study the mechanism of direct binding, oligomerization, and TIR domain activation of TNLs.
Abstract: INTRODUCTION Plants and animals respond to pathogen invasion through intracellular nucleotide-binding leucine-rich repeat receptors (NLRs) that directly interact with pathogen proteins or indirectly detect pathogen-derived alterations in the host proteome. Upon recognition of pathogen invasion, NLRs trigger an immune response that resolves in a variety of ways depending on the type of NLR being activated. The overall architecture of NLRs is highly conserved, consisting of a C-terminal leucine-rich repeat (LRR) platform that determines substrate specificity and a central nucleotide-binding oligomerization domain. The N-terminal domain varies between NLRs and determines the mechanism used by the host to activate the immune response. Thus, NLRs in plants have been classified according to their N-terminal domain into Toll/interleukin 1 receptor (TIR) NLRs (TNLs), coiled-coil NLRs (CNLs), and RPW8-like coiled-coil NLRs (RNLs). Pathogen detection and oligomerization of the NLR activates these N-terminal domains by bringing them in close contact. In all three cases, association of the N-terminal domain leads to localized cell death and expression of disease resistance. The TIR domains of TNLs have been shown to have oligomerization-dependent NADase activity that is required for promoting cell death, but it is not understood how the interactions between TIR domains renders them catalytically active. RATIONALE The structure of the ROQ1 (recognition of XopQ 1)–XopQ (Xanthomonas outer protein Q) complex, an immune receptor bound to its pathogen substrate, was used as a model to study the mechanism of direct binding, oligomerization, and TIR domain activation of TNLs. ROQ1 has been shown to physically interact with the Xanthomonas effector XopQ, causing it to oligomerize and trigger a TIR-dependent hypersensitive cell death response. We coexpressed, extracted, and purified the assembled ROQ1-XopQ complex from ROQ1’s native host, Nicotiana benthamiana, and solved its structure by cryo–electron microscopy to 3.8-A resolution. The interactions described in our structure were further confirmed by in vivo mutational analysis. RESULTS Our structure reveals that ROQ1 forms a tetrameric resistosome upon recognizing XopQ. The LRR and a post-LRR domain named the C-terminal jelly-roll/Ig-like domain (C-JID), form a horseshoe-shaped scaffold that curls around the pathogen effector, thereby recognizing multiple regions of the substrate. Binding of the ROQ1 LRR to XopQ occurs through surface-exposed residues that make up the scaffold of the domain, as well as an elongated loop between two LRRs that forms a small amphipathic α-helix at the site of interaction. The mode of substrate recognition by the C-JID is reminiscent of that used by immunoglobulins to bind their antigen. Similar to the complementary-determining regions of antibodies, interconnecting loops emerging from the C-JID β-sandwich structure make substrate-specific contacts with XopQ. In particular, an extended loop of the C-JID dives into the active-site cleft of XopQ and interacts with conserved residues required for nucleoside binding, suggesting that ROQ1 not only recognizes its substrate but also inhibits its ligand-binding function. The nucleotide-binding domain (NBD), helical domain 1 (HD1) and the winged-helix domain (WHD), termed NB-ARC because of their presence in Apaf-1, R proteins, and CED-4 (ARC), are responsible for ROQ1 oligomerization in an ATP-bound state. Individual protomers intercalate in a similar fashion as found in other NLR structures, promoting association between the N-terminal TIR domains. The TIR domains bind to each other through two distinct interfaces (called AE and BE), causing them to form a dimer of dimers. BE-interface contacts cause a conformational rearrangement in a loop, called the BB-loop, at the periphery of the TIR domain active site that exposes the putative catalytic glutamate that is suggested to cleave NAD+. These results provide a rationale for the previously determined oligomerization dependence of TIR domain NADase activity. CONCLUSION We propose a step-by-step mechanism for ROQ1 immune signaling based on our structure of the activated complex and on previous biochemical studies. The LRR and C-JID of ROQ1 recognize the pathogen effector through direct contacts with its surface and active-site residues. Detection of the substrate releases autoinhibitory contacts between the NB-ARC domain and the LRR, allowing the NB-ARC domain to transition to an ATP-bound, oligomerization-prone state. Complex assembly brings the TIR domains in close contact, leading to opening of the NADase active site in an interface-dependent manner. Cleavage of NAD+ by the TIR domain results in the release of adenosine diphosphate ribose, a signaling molecule that triggers cytosolic Ca2+ influx, a widely used chemical cue in response to various biotic and abiotic stresses, leading to downstream activation of localized cell death and disease resistance.

233 citations