skip to main content

Explore Research Products in the PAR
It may take a few hours for recently added research products to appear in PAR search results.


Title: Plasma membrane association and resistosome formation of plant helper immune receptors

Intracellular plant immune receptors, termed NLRs (Nucleotide-binding Leucine-rich repeat Receptors), confer effector-triggered immunity. Sensor NLRs are responsible for pathogen effector recognition. Helper NLRs function downstream of sensor NLRs to transduce signaling and induce cell death and immunity. Activation of sensor NLRs that contain TIR (Toll/interleukin-1receptor) domains generates small molecules that induce an association between a downstream heterodimer signalosome of EDS1 (EnhancedDisease Susceptibility 1)/SAG101 (Senescence-AssociatedGene 101) and the helper NLR of NRG1 (NRequired Gene 1). Autoactive NRG1s oligomerize and form calcium signaling channels largely localized at the plasma membrane (PM). The molecular mechanisms of helper NLR PM association and effector-induced NRG1 oligomerization are not well characterized. We demonstrate that helper NLRs require positively charged residues in their N-terminal domains for phospholipid binding and PM association before and after activation, despite oligomerization and conformational changes that accompany activation. We demonstrate that effector activation of a TIR-containing sensor NLR induces NRG1 oligomerization at the PM and that the cytoplasmic pool of EDS1/SAG101 is critical for cell death function. EDS1/SAG101 cannot be detected in the oligomerized NRG1 resistosome, suggesting that additional unknown triggers might be required to induce the dissociation of EDS1/SAG101 from the previously described NRG1/EDS1/SAG101 heterotrimer before subsequent NRG1 oligomerization. Alternatively, the conformational changes resulting from NRG1 oligomerization abrogate the interface for EDS1/SAG101 association. Our data provide observations regarding dynamic PM association during helper NLR activation and underpin an updated model for effector-induced NRG1 resistosome formation.

  more » « less
Award ID(s):
1758400
PAR ID:
10529144
Author(s) / Creator(s):
; ; ; ; ; ; ; ;
Publisher / Repository:
National Academy of Sciences
Date Published:
Journal Name:
Proceedings of the National Academy of Sciences
Volume:
120
Issue:
32
ISSN:
0027-8424
Format(s):
Medium: X
Sponsoring Org:
National Science Foundation
More Like this
  1. ; ; ; ; ; ; ; ; ; ; et al ( , Science)
    Plant nucleotide-binding leucine-rich repeat (NLR) immune receptors activate cell death and confer disease resistance by unknown mechanisms. We demonstrate that plant Toll/interleukin-1 receptor (TIR) domains of NLRs are enzymes capable of degrading nicotinamide adenine dinucleotide in its oxidized form (NAD + ). Both cell death induction and NAD + cleavage activity of plant TIR domains require known self-association interfaces and a putative catalytic glutamic acid that is conserved in both bacterial TIR NAD + -cleaving enzymes (NADases) and the mammalian SARM1 (sterile alpha and TIR motif containing 1) NADase. We identify a variant of cyclic adenosine diphosphate ribose as a biomarker of TIR enzymatic activity. TIR enzymatic activity is induced by pathogen recognition and functions upstream of the genes enhanced disease susceptibility 1 ( EDS1 ) and N requirement gene 1 ( NRG1 ), which encode regulators required for TIR immune function. Thus, plant TIR-NLR receptors require NADase function to transduce recognition of pathogens into a cell death response. 
    more » « less
  2. ; ; ; ; ; ; ; ; ; ; et al ( , New Phytologist)
    Summary

    Activation of nucleotide‐binding leucine‐rich repeat receptors (NLRs) results in immunity and a localized cell death. NLR cell death activity requires oligomerization and in some cases plasma membrane (PM) localization. The exact mechanisms underlying PM localization of NLRs lacking predicted transmembrane domains or recognizable lipidation motifs remain elusive.

    We used confocal microscopy, genetically encoded molecular tools and protein‐lipid overlay assays to determine whether PM localization of members of the Arabidopsis HeLo‐/RPW8‐like domain ‘helper’ NLR (RNL) family is mediated by the interaction with negatively charged phospholipids of the PM.

    Our results show that PM localization and stability of some RNLs and one CC‐type NLR (CNL) depend on the direct interaction with PM phospholipids. Depletion of phosphatidylinositol‐4‐phosphate from the PM led to a mis‐localization of the analysed NLRs and consequently inhibited their cell death activity. We further demonstrate homo‐ and hetero‐association of members of the RNL family. Our results provide new insights into the molecular mechanism of NLR localization and defines an important role of phospholipids for CNL and RNL PM localization and consequently, for their function.

    We propose that RNLs interact with anionic PM phospholipids and that RNL‐mediated cell death and immune responses happen at the PM.

     
    more » « less
  3. ; ; ; ; ; ; ; ; ; ; et al ( , Science)

    Plant nucleotide-binding leucine-rich repeat receptors (NLRs) regulate immunity and cell death. InArabidopsis, a subfamily of “helper” NLRs is required by many “sensor” NLRs. Active NRG1.1 oligomerized, was enriched in plasma membrane puncta, and conferred cytoplasmic calcium ion (Ca2+) influx in plant and human cells. NRG1.1-dependent Ca2+influx and cell death were sensitive to Ca2+channel blockers and were suppressed by mutations affecting oligomerization or plasma membrane enrichment. Ca2+influx and cell death mediated by NRG1.1 and ACTIVATED DISEASE RESISTANCE 1 (ADR1), another helper NLR, required conserved negatively charged N-terminal residues. Whole-cell voltage-clamp recordings demonstrated thatArabidopsishelper NLRs form Ca2+-permeable cation channels to directly regulate cytoplasmic Ca2+levels and consequent cell death. Thus, helper NLRs transduce cell death signals directly.

     
    more » « less
  4. ; ; ; ; ; ( , The Plant Journal)
    SUMMARY

    Plant intracellular immune receptors, primarily nucleotide‐binding, leucine‐rich repeat proteins (NLRs), detect pathogen effector proteins and activate NLR‐triggered immunity (NTI). Recently, ‘sensor’ NLRs have been reported to function with ‘helper’ NLRs to activate immunity. We investigated the role of two helper NLRs, Nrc2 and Nrc3, on immunity in tomato to the bacterial pathogenPseudomonas syringaepv. tomato (Pst) mediated by the sensor NLR Prf and the Pto kinase. Annrc2/nrc3mutant no longer activated Prf/Pto‐mediated NTI toPstcontaining the effectors AvrPto and AvrPtoB. Annrc3mutant showed intermediate susceptibility between wild‐type plants and aPrfmutant, while annrc2mutant developed only mild disease. These observations indicate that Nrc2 and Nrc3 act additively in Prf‐/Pto‐mediated immunity. We examined at what point Nrc2 and Nrc3 act in the Prf/Pto‐mediated immune response. In thenrc2/3mutant, programmed cell death (PCD) normally induced by constitutively active variants of AvrPtoB, Pto, or Prf was abolished, but that induced by M3Kα or Mkk2 was not. PCD induced by a constitutively active Nrc3 was also abolished in aNicotiana benthamianaline with reduced expression ofPrf. MAPK activation triggered by expression of AvrPto in the wild‐type tomato plants was completely abolished in thenrc2/3 mutant. These results indicate that Nrc2 and Nrc3 act with Prf/Pto and upstream of MAPK signaling. Nrc2 and Nrc3 were not required for PCD triggered by Ptr1, another sensor NLR‐mediatingPstresistance, although these helper NLRs do appear to be involved in resistance to certainPstrace 1 strains

     
    more » « less
  5. ; ; ; ; ; ; ; ; ; ; et al ( , Proceedings of the National Academy of Sciences)
    TIR domains are NAD-degrading enzymes that function during immune signaling in prokaryotes, plants, and animals. In plants, most TIR domains are incorporated into intracellular immune receptors termed TNLs. In Arabidopsis, TIR-derived small molecules bind and activate EDS1 heterodimers, which in turn activate RNLs, a class of cation channel–forming immune receptors. RNL activation drives cytoplasmic Ca 2+ influx, transcriptional reprogramming, pathogen resistance, and host cell death. We screened for mutants that suppress an RNL activation mimic allele and identified a TNL, SADR1. Despite being required for the function of an autoactivated RNL, SADR1 is not required for defense signaling triggered by other tested TNLs. SADR1 is required for defense signaling initiated by some transmembrane pattern recognition receptors and contributes to the unbridled spread of cell death in lesion simulating disease 1 . Together with RNLs, SADR1 regulates defense gene expression at infection site borders, likely in a non-cell autonomous manner. RNL mutants that cannot sustain this pattern of gene expression are unable to prevent disease spread beyond localized infection sites, suggesting that this pattern corresponds to a pathogen containment mechanism. SADR1 potentiates RNL-driven immune signaling not only through the activation of EDS1 but also partially independently of EDS1. We studied EDS1-independent TIR function using nicotinamide, an NADase inhibitor. Nicotinamide decreased defense induction from transmembrane pattern recognition receptors and decreased calcium influx, pathogen growth restriction, and host cell death following intracellular immune receptor activation. We demonstrate that TIR domains can potentiate calcium influx and defense and are thus broadly required for Arabidopsis immunity. 
    more » « less
  • Citation Formats
  • MLA
  • APA
  • Chicago
  • BibTeX
  • Save / Share this Record
  • Send to Email