Plant NLR diversity: the known unknowns of pan-NLRomes

Abstract

Plants and pathogens constantly adapt to each other. As a consequence, many members of the plant immune system, and especially the intracellular nucleotide-binding site leucine-rich repeat receptors, also known as NOD-like receptors (NLRs), are highly diversified, both among family members in the same genome, and between individuals in the same species. While this diversity has long been appreciated, its true extent has remained unknown. With pan-genome and pan-NLRome studies becoming more and more comprehensive, our knowledge of NLR sequence diversity is growing rapidly, and pan-NLRomes provide powerful platforms for assigning function to NLRs. These efforts are an important step toward the goal of comprehensively predicting from sequence alone whether an NLR provides disease resistance, and if so, to which pathogens.

Figure 1

Plant phylogeny and NLR complement sizes. Adapted from Gao et al. (2018), Munch et al. (2018), and Baggs et al. (2020).

Figure 2

Examples of possible changes in sequence and genomic organization in single-gene and clustered NLRs through evolution. Major changes can occur in an NLR locus, and often the reconstruction of the sequence of events will be impossible.

Figure 3

(A) Examples of allelic variation at individual NLR loci. P/A, presence/absence. R/S, resistant/susceptible. (B) RPP4/RPP5 NLR cluster across eight different A. thaliana accessions. Tall rectangles represent NLR genes, and short rectangles non-NLR genes. Colors indicate close sequence relatedness. There is a highly variable number of blue NLR genes in the cluster (including the functionally defined members RPP4, RPP5, and SNC1), while the adjacent RLM3 gene (magenta) shows P/A polymorphism. The RPP4/RPP5 cluster has also been invaded by an unrelated NLR gene (yellow). Finally, some of the non-NLR genes (pink, green) flanking the cluster are duplicated as well. (C) Average number of genes per NLR gene cluster across eight A. thaliana accessions. Most have on average fewer than three genes, while a few are both highly expanded and highly variable in numbers. Data for B and C from Jiao and Schneeberger (2020).

Figure 4

Potential workflow of the construction, visualization, and analysis of a pan-NLRome. First, accessions to be sequenced are selected in a manner that maximizes diversity, in order to approach a saturated pan-NLRome as quickly as possible (1). Then either the whole genome or the NLR complement is sequenced and assembled (2). The assemblies from each genome are then aligned to each other and potentially combined into a genome graph (3). Core and accessory genes are identified and the saturation point of the NLRome is assessed, to determine whether the NLRome is open or closed (4). Finally, individual NLR groups can be studied, e.g. one can analyze the phylogeny of a certain cluster across accessions, by comparing individual alleles in those accessions that share the same genes, the structure of whole clusters, or the entire set of genes present in all clusters (5).