The self-association of Toll/interleukin-1 receptor/resistance protein (TIR) domains has been implicated in signaling in plant and animal immunity receptors. Structure-based studies identified different TIRdomain dimerization interfaces required for signaling of the plant nucleotide-binding oligomerization domain-like receptors (NLRs) L6 from flax and disease resistance protein RPS4 from Arabidopsis. Here we show that the crystal structure of the TIR domain from the Arabidopsis NLR suppressor of npr1-1, constitutive 1 (SNC1) contains both an L6-like interface involving helices αD and αE (DE interface) and an RPS4-like interface involving helices αA and αE (AE interface). Mutations in either the AE-or DE-interface region disrupt cell-death signaling activity of SNC1, L6, and RPS4 TIR domains and full-length L6 and RPS4. Selfassociation of L6 and RPS4 TIR domains is affected by mutations in either region, whereas only AE-interface mutations affect SNC1 TIR-domain self-association. We further show two similar interfaces in the crystal structure of the TIR domain from the Arabidopsis NLR recognition of Peronospora parasitica 1 (RPP1). These data demonstrate that both the AE and DE self-association interfaces are simultaneously required for self-association and cell-death signaling in diverse plant NLRs.plant immunity | NLR | TIR domain | plant disease resistance | signaling by cooperative assembly formation P lants have evolved a sophisticated innate immune system to detect pathogens, in which plant resistance (R) proteins recognize pathogen proteins (effectors) in a highly specific manner. This recognition leads to the effector-triggered immunity (ETI) response that often induces a localized cell death known as the hypersensitive response (1). Most R proteins belong to the nucleotide-binding oligomerization domain (NOD)-like receptor (NLR) family. NLRs are prevalent in the immune systems of plants and animals and provide resistance to a broad range of pathogens, including fungi, oomycetes, bacteria, viruses, and insects (2, 3). NLRs contain a central nucleotide-binding (NB) domain, often referred to as the nucleotide-binding adaptor shared by APAF-1, resistance proteins, and CED-4 (NB-ARC domain) (4) and a C-terminal leucine-rich repeat (LRR) domain. Plant NLRs can be further classified into two main subfamilies, depending on the presence of either a Toll/interleukin-1 receptor domain (TIR-NLR) or a coiled-coil domain (CC-NLR) at their N termini (5).The CC and TIR domains of many plant NLRs can autonomously signal cell-death responses when expressed ectopically in planta, and mutations in these domains within full-length proteins also compromise signaling, suggesting that these domains are responsible for propagating the resistance signal after activation of the receptor (6-14). Self-association of both TIR (8, 9, 11, 15) and CC (10,13,16,17) domains has been shown to be important for the signaling function. In animal NLRs, the formation of postactivation oligomeric complexes, such as the NLRC4/NAIP inflammasome or...

Plant nucleotide-binding leucine-rich repeat (NB-LRR) disease resistance (R) proteins recognize specific “avirulent” pathogen effectors and activate immune responses. NB-LRR proteins structurally and functionally resemble mammalian Nod-like receptors (NLRs). How NB-LRR and NLR proteins activate defense is poorly understood. The divergently transcribed Arabidopsis R genes, RPS4 (resistance to Pseudomonas syringae 4) and RRS1 (resistance to Ralstonia solanacearum 1), function together to confer recognition of Pseudomonas AvrRps4 and Ralstonia PopP2. RRS1 is the only known recessive NB-LRR R gene and encodes a WRKY DNA binding domain, prompting suggestions that it acts downstream of RPS4 for transcriptional activation of defense genes. We define here the early RRS1-dependent transcriptional changes upon delivery of PopP2 via Pseudomonas type III secretion. The Arabidopsis slh1 (sensitive to low humidity 1) mutant encodes an RRS1 allele (RRS1SLH1) with a single amino acid (leucine) insertion in the WRKY DNA-binding domain. Its poor growth due to constitutive defense activation is rescued at higher temperature. Transcription profiling data indicate that RRS1SLH1-mediated defense activation overlaps substantially with AvrRps4- and PopP2-regulated responses. To better understand the genetic basis of RPS4/RRS1-dependent immunity, we performed a genetic screen to identify suppressor of slh1 immunity (sushi) mutants. We show that many sushi mutants carry mutations in RPS4, suggesting that RPS4 acts downstream or in a complex with RRS1. Interestingly, several mutations were identified in a domain C-terminal to the RPS4 LRR domain. Using an Agrobacterium-mediated transient assay system, we demonstrate that the P-loop motif of RPS4 but not of RRS1SLH1 is required for RRS1SLH1 function. We also recapitulate the dominant suppression of RRS1SLH1 defense activation by wild type RRS1 and show this suppression requires an intact RRS1 P-loop. These analyses of RRS1SLH1 shed new light on mechanisms by which NB-LRR protein pairs activate defense signaling, or are held inactive in the absence of a pathogen effector.

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Some plant pathogens can infect an extremely diverse range of hosts (Newman & Derbyshire, 2020). Among these is the necrotrophic fungus Sclerotinia sclerotiorum, which stands out for its ability to cause significant disease in numerous crops. Disease caused by this pathogen is given various names, though one of the most common seems to be sclerotinia stem rot. S. sclerotiorum infects crops in almost all agricultural production zones ranging from dry bean (Phaseolus vulgaris) in the tropics (Lehner et al., 2017) to canola (Brassica napus