Ligand Recognition and Immune Activation by Plant NLR Receptor Pairs – RePairs
The goal of the RePairs project is to generate fundamental and transferable knowledge on plant immune receptors from the family of nucleotide-binding (NB) and leucine-rich repeat domain (LRR) proteins (NLRs). NLRs form a large family of intracellular receptors that detect virulence factors (also called effector proteins) secreted by pathogens to promote disease and that trigger immune reactions in the host cell to prevent the spread of infection.
NLRs are the most important class of plant disease resistance proteins that render crops resistant to major pathogens. Introducing them into elite varieties by breeding or genetic engineering represents an economically and ecologically outstandingly advantageous solution for plant disease control2. NLRs have therefore a critical role for sustainable crop disease management.
Until recently, the mechanistic understanding of NLRs has been hampered by the lack of suitable in vitro systems and limited structural knowledge. Breakthroughs in the purification of NLRs from eukaryotic expression systems and in protein structure determination by cryo-electron microscopy (cryo-EM) are lifting these barriers. They revealed that plant NLRs, like other signaling ATPases from the STAND protein family assemble upon effector recognition in signaling competent oligomeric rings called resistosomes. The RePairs project builds on these technical and conceptual advances to produce new fundamental discoveries in NLR biology and plant immune research.
RePairs will focus on heterologous NLR pairs consisting of one sensor NLR (sNLR) that detects specific effectors from pathogens, and one executor NLR (eNLR) that initiates immune signalling. These paired NLRs are still poorly understood but provide outstanding perspectives for the engineering of resistance proteins with novel specificities.
Our central hypothesis is that effector binding to multiple sites in the sNLR induces conformational changes in the inactive sNLR/eNLR complex that triggers assembly of an active oligomeric resistosome. So far, the oligomeric status, composition and three-dimensional rearrangement of such complexes remain elusive in case of NLR pairs.
To test this hypothesis we will elucidate the molecular mechanisms of effector recognition by sNLRs (Objective 1) and decipher how effector recognition activates sNLR/eNLR complexes (Objective 2). Our aim is then to employ the molecular and mechanistic insight generated by this research to engineer NLR hetero pairs with new recognition specificities (objective 3). Finally, we will investigate how NLR pairs acquire novel recognition specificities during evolution and how sNLR and eNLRs co-evolve (objective 4).
As an experimental system we will use the rice NLR pair RGA4/RGA5 that we have established as a leading model for the understanding of sNLR/eNLR pairs. Our previous studies as well as extensive preliminary data and biological resources for this model system will ensure success and rapid progress in the research tasks.
The project brings together a strong and diverse research consortium to deploy a highly interdisciplinary and integrative approach for a better understanding of NLRs. Major activities will be in vitro studies with cutting edge approaches such as native Mass Spectrometry (MS) and the structural characterization of NLRs and their complexes using hydrogen deuterium exchange (HDX) MS and cryo-EM. Insight from these analyses will be challenged by in vivo experiments using cellular and whole plant systems. These experimental approaches will be completed by comparative genomics and modelling of 3D structures of protein and their complexes.
At the completion of this work, the RePairs project will provide major advances in plant immune research by revealing the structure and molecular mechanism of sNLR/eNLR pairs, elucidating the coevolution of paired NLRs and generating novel effector recognition specificities by molecular engineering of NLR immune receptor pairs.
Monsieur Thomas KROJ (Plant Health Institute Montpellier)
The author of this summary is the project coordinator, who is responsible for the content of this summary. The ANR declines any responsibility as for its contents.
IGF Institut de génomique fonctionnelle
KAUST Stefan T Arold
PHIM Plant Health Institute Montpellier
CBS Centre de biochimie structurale
Help of the ANR 575,311 euros
Beginning and duration of the scientific project: December 2021 - 48 Months