Two NLR immune receptors acquired high-affinity binding to a fungal effector through convergent evolution of their integrated domain

Abstract

A subset of plant NLR immune receptors carry unconventional integrated domains in addition to their canonical domain architecture. One example is rice Pik-1 that comprises an integrated heavy metal–associated (HMA) domain. Here, we reconstructed the evolutionary history of Pik-1 and its NLR partner, Pik-2, and tested hypotheses about adaptive evolution of the HMA domain. Phylogenetic analyses revealed that the HMA domain integrated into Pik-1 before Oryzinae speciation over 15 million years ago and has been under diversifying selection. Ancestral sequence reconstruction coupled with functional studies showed that two Pik-1 allelic variants independently evolved from a weakly binding ancestral state to high-affinity binding of the blast fungus effector AVR-PikD. We conclude that for most of its evolutionary history the Pik-1 HMA domain did not sense AVR-PikD, and that different Pik-1 receptors have recently evolved through distinct biochemical paths to produce similar phenotypic outcomes. These findings highlight the dynamic nature of the evolutionary mechanisms underpinning NLR adaptation to plant pathogens.

The allelic rice immune receptor Pikh confers extended resistance to strains of the blast fungus through a single polymorphism in the effector binding interface

“…However, the Asn261 polymorphism is present in Pik orthologs from some wild rice species that diverged from cultivated rice [54]. Additionally, in-depth evolutionary analysis and ancestral gene reconstruction of Pik-HMA domains also places Asn261 as the ancestral residue at this position [55]. This further supports the hypothesis that Pikm and Pikh appear to have convergently evolved towards the same molecular mechanism to extend effector recognition specificity, by having a lysine residue one position towards the N-terminus in their protein sequence, compared with the narrow-spectrum allele Pikp.…”

Functional diversification gave rise to allelic specialization in a rice NLR immune receptor pair

“…The rice NLRs Pik-1 and Pik-2 form a linked gene pair arranged in an inverted configuration on chromosome 11 with only ∼2.5 Kb separating their start codons (Ashikawa et al, 2008). The Pik pair is present in the genetic pool of rice cultivars as two major haplotypes (Bialas et al, 2021; Kanzaki et al, 2012). Pik pairs belonging to the K haplotype confer resistance to strains of the rice blast fungus, Magnaporthe oryzae , that harbour the effector AVR-Pik (Ashikawa et al, 2008).…”

“…Allelic Pik NLRs have differential recognition specificities for the AVR-Pik variants (De la Concepcion et al, 2021; Kanzaki et al, 2012), which is underpinned by differential effector binding to the Pik-1 HMA domain (De la Concepcion et al, 2018; De la Concepcion et al, 2021; Maqbool et al, 2015). Two allelic variants of Pik-1, Pikp-1 and Pikm-1, acquired high-affinity binding to the M. oryzae AVR-Pik effector through convergent evolution of their HMA domains (Bialas et al, 2021). Additionally, Pikm-1 and Pikh-1 alleles have been shown to convergently evolve towards extended recognition specificity of AVR-Pik variants (De la Concepcion et al, 2021).…”

“…Additionally, Pikm-1 and Pikh-1 alleles have been shown to convergently evolve towards extended recognition specificity of AVR-Pik variants (De la Concepcion et al, 2021). This adaptive evolution towards recognition of rapidly evolving effectors has led to marked diversification of the integrated HMA domain (Bialas et al, 2021). As a consequence, Pik-1 HMA domain is the most sequence-diverged domain in the Pik NLR pair (Bialas et al, 2021; Bialas et al, 2018; Costanzo and Jia, 2010).…”

“…While Pik-1 acts as a sensor, Pik-2 acts as a helper NLR that is required for the activation of immune responses (Maqbool et al, 2015; Zdrzalek et al, 2020). Evolutionary analyses have shown that the genetic linkage of this NLR pair is ancient and revealed marked signatures of adaptive evolution in the integrated HMA domain of Pik-1 (Bialas et al, 2021). However, little is known about sensor/helper coevolution in Pik and how these multidomain proteins have adapted to changes in the rapidly evolving integrated HMA domain of Pik-1.…”