8. VL Spallek ETI

• effectors are proteins produced by pathogens (fungi, bacteria) to attack the plant (and verse visa?)

• if they are recognized by plant they activate the immune response of the plants

• an effector that is recognized by a R protein is called Avr (=avirulence) protein

• apoplast: PRRs

• intracellular: NLR (intracellular nucleotide-binding leucine-rich repeat receptors)

NLR = Nucleotide binding Leucine rich-repeat Receptors

• NLRs detect R gene-mediated resistance (also called gene-for-gene resistance, effector triggered immunity (ETI))

• basic structure: LRR-domain (leucine rich repeat) for protein-protein interactions (conserved protein fold)

  • deletion of LRRs activate NLR signaling = negative regulation of NBD-mediated oligomerization

• basic structure: NB-domain (Nucleotide binding) —> binds and hydrolyzes ATP (on/off switch)

Major protein classes:

• TNLs (Toll/Interleukin-1 Receptors domain (TIR) NLRs

  • lost in monocots

• CNLs (coiled-coil domain (CC) NLRs)

  • subdomain: RNLs (RPW8-like CC-domain NLR) = helper NLRs

singleton = active without additional R proteins

singleton = active without additional R proteins

1. AvrAC = Effector —> uridylates PBL2 (=RLCK) —> UMP is recognized by ZAR1 (=singleton CNL)

   RKS1 = pseudokinase

2. formation of ZAR1/RKS heterodimer, association with uridylated PBL2UMP —> confirmation change —> ADP of ZAR1 is ejected and replaced by ATP = ACTIVATION —> oligomerization of ZAR1/RKS2/PBL2UMP = ZAR1-resistome (pentamer)

3. ZAR1-resistome localizes to plasma membrane

4. 5. 6. resistome on plasmamembrane triggers Ca2+ channel activity —> Ca2+ influx; ROS-accumulation; activation of downstream signaling

   1. Upregulation of defense genes

   2. 9. 10. production of defense-related phytohormones (e.g. SA); pertubation of chloroplast and vacuoles; loss of plasma membrane intergrity = unset of HR

Zu genau? Zusammenfassung eine Folie danach:

1. RPP1 = TNL; ATR = effector from H. arabidopsis

   RPP1 binds ATR1

2. binding leads to conformational changes and oligomerization tetramer —> TNL resistome (TIR domain in close proximity)

3. close proximity of TIR domains trigger NADase and nuclease activity

   production of small signaling molecules: v-cADPR (=cycle adenosine diphosphate ribose) + 2’3’-cAMP production

4. activation of EP proteins

   complex formation with helper NLRs: EDS1+PAD4+ADR1 & EDS1+SAG101+NRG1

5. + 8. helper NLRs form Ca2+ channels, induction of HR

6. + 7. activation of TFs and transcriptional reprogramming by EP complexes and ultimately in biosynthesis of defense compounds and desfense-related phytohormones

Zu genau? Zusammenfassung eine Folie danach:

• required NLRs

• oligomerization through N-terminal domains

HR = hypersensitive response

—> rapid localized cell death that occurs at the point of pathogen penetration and is associates with disease resistance

—> induced via NLR signaling

similarities: release of DAMPs (=Danger-Associates Molecular Patterns)

differences: execution of cell death (in animals necroptosis, ferroptosis, apoptosis)

biotrophic: pathogen keeps host alive

necrotrophic: pathogen kills host

1. (Direct interaction)

1. Guard hypothesis

   • example: Arabidopsis RPS2 <—> Pseudomonas AvrRpt2

2. Zig-Zag-model

   • example: FLS2-AvrPtoB-Pto

3. Decoy model

   • FLS2-AvrPtoB-Pto (Pto/Fen is decoy), ZAR1 (PBL2 is decoy), RPS5 (PBS1/PBL2 is decoy)

4. Integrated decoy model

1. Direct interaction

   • R protein is a receptor and effector is the ligand

   • only few examples support this model and hard to explain how the relatively small R protein repertoire can target the broad diversity of plant pathogens

   • 
     

Alternative hypothesis “guard hypothesis”:

• R proteins monitor (“guard”) the integrity of host cellular targets of effector action —> such indirect detection of pathogen allows a limited number of R proteins to detect the activity of multiple pathogen effectors targeting points of vulnerability

• key tenets of hypothesis:

  • effector acting as a virulence factor has target(s) in host

  • by manipulating/ altering target(s) the effector contributes to pathogen success in susceptible host genotypes

  • effector perturbation of host target generates ‘pathogen-induced modified self’ molecular pattern which activates the corresponding R protein leading to ETI

example: Arabidopsis RPS2 <—> Pseudomonas AvrRpt2

• RPS2 detects AvrRpt2-mediated RIN4 cleavage

• AvrRpt2 is a protease that directly cleaves RIN4 (=PM-associated regulator of basal defense)

1. PAMPs are recognized by their cognate PRRs in plants leading to PTI

2. Pathogen develops effectors that disable PAMP recognition leading to effector-triggered susceptibility (ETS)

3. Effectors become aivirulence factors (Avrs) once the plant developed cognate resistance protein (R-protein) leading to ETI

4. Pathogen develops new effectors and/or loses its Avrs to circumvent ETI again leading to ETS which then repeatedly brings the plant to develop new R-proteins leading to ETI

Example: FLS2-AvrPtoB-Pto

• AvrPtoB = Pseudomonas effector that binds kinases of PRRs

• evolutionarily, guarded effector target is an unstable situation (subject to 2 opposing natural selection forces: absence R gene: natural selection drives guardee to decrease binding affinity with effector; presence R gene: natural selection favors guardee with improved interaction with an effector to enhance pathogen perception)

• evolutionarily unstable situation can be relaxed upon evolution of host protein: decoy

• decoy specializes in perception of effector by the R protein but itself has no function either in development of disease or resistance

• decoy mimics effector targets to trap the pathogen into a recognition event

examples: FLS2-AvrPtoB-Pto (Pto/Fen is decoy), ZAR1 (PBL2 is decoy), RPS5 (PBS1/PBL2 is decoy)

• plant protein targeted by an effector has been duplicated and fused to one member of the NLR pair

• there, it acts as a bait to trigger defense signaling by the second NLR upon effector binding

• examples: AvrRps4/PopP2 —> inhibit RRS1 WRKY domain —> inhibit RRS1/PRS4

• Example (RRS1-R)

  RRS1-R (R protein) has a WRKY domain, which can be bound by effectors (e.g., AvrRps4 and PopP2), leading to the binding of the WRKY domain to W-boxed (=> cis-regulatory elements of many defense genes)

  RRS1-R has an additional helper NLR; without it, RRS1-R is inactive; RRS1-R is the sensor NLR

  Sensor NLR and helper NLR are often organized in clusters with head-to-head orientation 

• ZAR1: oligomer of NLR

• in CNL-ZAR1 signaling

• localizes on cytosolic site of plasma membrane to mediate Ca2+ influx

  
  

• for each gene controlling resistance (R-gene) in the host, there is a corresponding, specific gene controlling avirulence (Avr) gene in the pathogen

• has been observed between parasitic plants and their host (e.g. HaOr7 confers resistance to the parasitic plant O. cumana race F in sunflower)

• only when pathogen has Avr gene and the plant the R-gene —> recognition. In other cases no recognition

• EDS1 = Enhanced Disease Susceptibility 1

• essential component of R gene-mediated disease resistance

• form complexes with helper NLRs

• RPW8-like CC-domains (RNLs)

• form complexes with e.g. EDS1 to form a Ca2+ channel (TNL signaling + helper NLR)

Jessi:

• NAD+ hydrolysing enzymes that promote cell death?

• https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7045805/

Annabel: from lecture:

TIR domains oligomerize enzymes that produce small signaling molecules (EDS1/PAD4 and EDS1/SAG101)

• the activation of PTI is an indespensable component of ETI during bacterial infection

• in mutant plants in which the PTI signaling cascade is disabled, ETI is also impaired

• gene-for-gene model can’t be applied since multiple genes have to be present in order for the reaction to be incompatible

gene-for-gene resistance, race-specific resistance or Effector-triggered Immunity (ETI)