Plant Cell | Highly variable NLR immunoreceptor and prediction of its binding site with effector

Written by Jenny | Wang Yi In the immune system of plants, Nucleotide-Binding Leucine Rich Repeat (NLR) is an important type of immune recognition receptor, which can recognize pathogen effectors in the cell and cause hypersensitivity reactions in plants ( hypersensitive response, HR), resist the infection of pathogenic bacteria.

The specific evolution of the immune recognition system depends on the generation of receptor diversity, the acquisition of new antigen binding ability and the initiation of downstream signals.

However, this diversity may lead to mismatches between NLR and plant's own ligands during the hybridization process and initiate an immune response, thereby causing plant hybrid neucrosis [1].

A part of NLR can directly bind to the effector, while the other part activates the immune response by indirectly recognizing the modification of the plant protein by the effector.

However, how to determine the NLR that directly or indirectly binds to the effector according to the sequence, the relationship between the generation and evolution of NLR diversity within the same species is not clear [2].

As early as 1998, Michelmore and Meyers proposed the theory that in all NLRs, highly variable amino acid positions should correspond to their ligand binding sites [3].

But as more highly conservative, indirectly binding effector NLRs are discovered, this theory has been challenged.

Therefore, in order to clarify the correspondence between the highly variable amino acid sites of NLR and the ligand binding sites, it is particularly important to develop new methods to study the evolution of NLR and predict the binding sites of effectors.

Recently, Daniil M.
Prigozhin of Berkeley National Laboratory in the United States and Ksenia V Krasileva of the University of California at Berkeley jointly published a research paper entitled Analysis of intraspecies diversity reveals a subset of highly variable plant immune receptors and predicts their binding sites in The Plant Cell.
.

The study found highly variable NLRs by analyzing the sequence of NLR protein in Arabidopsis thaliana and Brachypodium dilatum, and predicted the binding sites of these NLRs and effectors.

In this study, in order to obtain NLR near allelic series (single copy orthogroups), the authors first constructed NLR genes of 62 different Arabidopsis ecotypes.
The phylogenetic tree is then divided into 65 branches according to the clade size and bootstrap support of the phylogenetic tree branches.

The author used the Shannon entropy method to analyze the sequence variability of each group of NLRs genes and found that all known NLRs that directly bind to effectors are highly variable, while all known indirect NLRs that bind to effectors are highly variable.
Conservative.

Further analysis of these highly variable NLRs shows that their variable amino acid residues are mainly distributed in the LRR domain that plays a role in antigen recognition.

Through the analysis of the known structure of RPP1, it is found that its effector binding sequence shows diversity.

These evidences indicate that the above methods can be used to determine whether a certain NLR can directly bind to the effector and predict its binding site.

Combining the position of highly variable and conservative NLR in the phylogenetic tree, the author proposes that the NLR that directly binds to effectors originated from the same ancestor, and the NLR that directly binds to effectors has undergone diversifying selection, and NLR that does not directly bind to effectors has undergone an evolutionary process of purifying or balancing selection.

Figure: RPP1 and effector binding sites show high sequence diversity.

In order to verify whether the conclusions obtained from the traceability analysis of different types of NLR using the phylogenetic tree method are universal in other species, the author also analyzed the NLR of 54 different ecotypes of Brachypodium difficile and found its evolution.
The trees showed a similar distribution, and 40 highly variable NLR genes were obtained.

The author also combined the reference genomes of Arabidopsis Col-0 and Brachypodium erbatarum Bd-21 and found that only a small number of genes from different genomes are closer, indicating that the NLR evolutionary relationship between species is farther.

In conclusion, this article found that the method of using evolutionary tree and Shannon entropy can be used to analyze the evolutionary relationship of NLR within species, and predict whether NLR can directly bind to effectors and the sites where they bind.

At the same time, these highly variable genes are likely to be the gene loci that cause hybridization incompatibility.
Predicting these gene loci can provide guidance for future agricultural breeding.

However, the method of finding near allelic series based on the phylogenetic tree proposed in this paper requires a lot of manual calculation.
This problem needs to be solved by improving the algorithm of phylogenetic tree analysis.References 1.
Bomblies, K.
(2009).
Too much of a good thing? Hybrid necrosis as a by-product of plantimmune system diversification.
Botany 87: 1013–1022.
2.
Cesari, S.
(2018).
Multiple strategies for pathogen perception by plant immune receptors.
NewPhytol.
219: 17–24.
3.
Michelmore, RW and Meyers, BC (1998).
Clusters of resistance genes in plants evolve by divergent selection and a birth-and-death process.
Genome Res.
8: 1113– 1130.
Link to the original text: https://academic.
oup.
com/plcell/advance-article/doi/10.
1093/plcell/koab013/6119334