NLR-Associating Transcription Factor bHLH84 and Its Paralogs Function Redundantly in Plant Immunity

PLoS Pathogens, Aug 2014

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.

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NLR-Associating Transcription Factor bHLH84 and Its Paralogs Function Redundantly in Plant Immunity

et al. (2014) NLR-Associating Transcription Factor bHLH84 and Its Paralogs Function Redundantly in Plant Immunity. PLoS Pathog 10(8): e1004312. doi:10.1371/journal.ppat.1004312 NLR-Associating Transcription Factor bHLH84 and Its Paralogs Function Redundantly in Plant Immunity Fang Xu 0 Paul Kapos 0 Yu Ti Cheng 0 Meng Li 0 Yuelin Zhang 0 Xin Li 0 Bart Thomma, Wageningen University, Netherlands 0 1 Michael Smith Laboratories, University of British Columbia , Vancouver, British Columbia , Canada , 2 Department of Botany, University of British Columbia , Vancouver, British Columbia , Canada , 3 National Institute of Biological Sciences , Beijing , People's Republic of China 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. - Funding: This research is supported from financial funds from the Natural Sciences and Engineering Research Council of Canada (NSERC) Discovery program and the William Cooper Endowment Fund from UBC Botany Department. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. Plants have evolved a sophisticated immune system to fight against invading microbial pathogens that threaten their normal growth and development. Plant immunity is in part mediated by resistance (R) proteins that recognize pathogen proteins known as effectors [13]. The majority of R proteins are NLR receptors that contain leucine-rich repeats (LRRs) at the C-terminus, a central nucleotide-binding site (NBS) and either a Toll/Interleukin-1 receptor (TIR) or a coiled-coil (CC) domain at the N-terminus [4]. In Arabidopsis, genetically downstream of the R proteins are the EDS1 (ENHANCED DISEASE SUSCEPTIBILITY 1)/PAD4 (PHYTOALEXIN DEFICIENT 4)/SAG101 (SENESCENCEASSOCIATED GENE101) complex and NDR1 (NON-RACERESISTANCE 1), which mainly mediate TIR-NB-LRR or CCNB-LRR triggered defense responses, respectively [58]. While the mechanisms underlying effector recognition by R proteins have been intensively studied, little is known about the post-recognition events leading to defense activation. Recently, it has been shown that the nuclear pool of certain R proteins, including MLA10 (MILDEW A LOCUS 10) in barley, N in tobacco, Pb1 (Panicle blast 1) in rice, and RPS4 (RESISTANT TO P.SYRINGAE 4), RRS1 (RESISTANT TO RALSTONIA SOLANACEARUM 1) and SNC1 (SUPPRESSOR OF NPR1-1, CONSTITUTIVE1) in Arabidopsis, is important for the activation of defense responses [914]. The latest discoveries on the interactions between some of these R proteins and their associating transcription factors (TFs) further shed light on the activation mechanism of nuclear R proteins. For example, MLA10 interacts with WRKY TFs to de-repress PAMP (PATHOGEN-ASSOCIATED MOLECULAR PATTERN) triggered basal defense [9]. The active state of MLA10 can also release MYB6 (MYB DOMAIN PROTEIN 6) from WRKY suppression and promote its binding to cis-elements to initiate defense responses [15]. CCtype NLR Pb1 in rice interacts with WRKY45 and this interaction is believed to protect the TF from proteasomal degradation in the nucleus [16]. In addition, SNC1 associates with transcriptional corepressor TPR1 (TOPLESS RELATED 1) to negatively regulate the expression of known defense suppressors, thereby activating plant immunity [17]. Lately, studies on N in tobacco showed that it (...truncated)


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Fang Xu, Paul Kapos, Yu Ti Cheng, Meng Li, Yuelin Zhang, Xin Li. NLR-Associating Transcription Factor bHLH84 and Its Paralogs Function Redundantly in Plant Immunity, PLoS Pathogens, 2014, 8, DOI: 10.1371/journal.ppat.1004312