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In plants and animals, nucleotide-binding and leucine-rich repeat domain containing (NLR) immune receptors are utilized to detect the presence or activities of pathogen-derived molecules. However, the mechanisms by which NLR proteins induce defense responses remain unclear. Here, we report the characterization of one basic Helix-loop-Helix (bHLH) type transcription factor (TF), bHLH84, identified from a reverse genetic screen. It functions as a transcriptional activator that enhances the autoimmunity of NLR mutant snc1 (suppressor of npr1-1, constitutive 1) and confers enhanced immunity in wild-type backgrounds when overexpressed. Simultaneously knocking out three closely related bHLH paralogs attenuates RPS4-mediated immunity and partially suppresses the autoimmune phenotypes of snc1, while overexpression of the other two close paralogs also renders strong autoimmunity, suggesting functional redundancy in the gene family. Intriguingly, the autoimmunity conferred by bHLH84 overexpression can be largely suppressed by the loss-of-function snc1-r1 mutation, suggesting that SNC1 is required for its proper function. In planta co-immunoprecipitation revealed interactions between not only bHLH84 and SNC1, but also bHLH84 and RPS4, indicating that bHLH84 associates with these NLRs. Together with previous finding that SNC1 associates with repressor TPR1 to repress negative regulators, we hypothesize that nuclear NLR proteins may interact with both transcriptional repressors and activators during immune responses, enabling potentially faster and more robust transcriptional reprogramming upon pathogen recognition., Author Summary In plants and animals, NLR immune receptors are utilized to detect pathogen-derived molecules and activate immunity. However, the mechanisms of plant NLR activation remain unclear. Here, we report on bHLH84, which functions as a transcriptional activator. Simultaneously knocking out three closely related bHLH paralogs partially suppresses the autoimmunity of snc1 and compromises RPS4-mediated defense, while overexpression of these close paralogs renders strong autoimmunity, suggesting functional redundancy in the gene family. In planta co-immunoprecipitation revealed interactions between not only bHLH84 and SNC1, but also bHLH84 and RPS4. Therefore bHLH84 family transcription factors associate with these NLRs to activate defense responses, enabling potentially faster and more robust transcriptional reprogramming upon pathogen recognition.

Plant defense responses need to be tightly regulated to prevent auto-immunity, which is detrimental to growth and development. To identify negative regulators of Resistance (R) protein-mediated resistance, we screened for mutants with constitutive defense responses in the npr1-1 background. Map-based cloning revealed that one of the mutant genes encodes a conserved TPR domain-containing protein previously known as SRFR1 (SUPPRESSOR OF rps4-RLD). The constitutive defense responses in the srfr1 mutants in Col-0 background are suppressed by mutations in SNC1, which encodes a TIR-NB-LRR (Toll Interleukin1 Receptor-Nucleotide Binding-Leu-Rich Repeat) R protein. Yeast two-hybrid screens identified SGT1a and SGT1b as interacting proteins of SRFR1. The interactions between SGT1 and SRFR1 were further confirmed by co-immunoprecipitation analysis. In srfr1 mutants, levels of multiple NB-LRR R proteins including SNC1, RPS2 and RPS4 are increased. Increased accumulation of SNC1 is also observed in the sgt1b mutant. Our data suggest that SRFR1 functions together with SGT1 to negatively regulate R protein accumulation, which is required for preventing auto-activation of plant immunity., Author Summary The nucleotide-binding domain and leucine-rich repeats-containing (NLR) proteins are structurally conserved immune receptors found in both animals and plants. Correct folding of NLR proteins requires two conserved proteins, SGT1 and HSP90. We showed that another evolutionarily conserved protein, SRFR1, interacts with SGT1 in both yeast two-hybrid assays and co-immunoprecipitation analysis. Loss-of-function mutations in SRFR1 result in constitutive activation of immune responses. The constitutive activation of immune responses in the srfr1 mutants is dependent on the NLR Resistance (R) protein SNC1. In srfr1 mutant plants, levels of multiple R proteins including SNC1, RPS2 and RPS4 are elevated. Consistent with previous findings that SGT1b is involved in the negative regulation of protein levels of certain NLR R proteins, increased accumulation of SNC1 is also observed in the sgt1b mutant. Our data suggest that SRFR1 functions together with SGT1 to negatively regulate NLR R protein accumulation to prevent autoimmunity in plants.