An unusual tandem kinase fusion protein confers leaf rust resistance in wheat

Abstract

The introgression of chromosome segments from wild relatives is an established strategy to enrich crop germplasm with novel disease resistance genes. Here, we use mutagenesis and transcriptome sequencing to clone the leaf rust resistance gene Lr9, which was introduced into bread wheat from the wild grass species Aegilops umbellulata. We establish that Lr9 encodes an unusual tandem kinase fusion protein. Long-read sequencing of a wheat Lr9 introgression line and the putative Ae. umbellulata Lr9 donor enabled us to assemble the ~28.4-Mb Lr9 translocation and to identify the translocation breakpoint. We likewise cloned Lr58, which was reportedly introgressed from Ae. triuncialis, but has an identical coding sequence compared to Lr9. Cytogenetic and haplotype analyses corroborate that the two genes originate from the same translocation event. Our work sheds light on the emerging role of kinase fusion proteins in wheat disease resistance, expanding the repertoire of disease resistance genes for breeding.

Resistance that stacks up: engineering rust and mildew disease control in the cereal crops wheat and barley

“…Moreover, unbiased complexity reduction approaches, such as TACCA (targeted chromosome‐based cloning via long‐range assembly) or MutChromSeq, that are based on chromosome sorting and sequencing of wild‐type plants and chemically induced mutants have been applied to isolating genes involved in disease resistance (Dracatos et al ., 2019; Sánchez‐Martín et al ., 2016; Thind et al ., 2017; Yu et al ., 2022) and morphological characteristics (Ford et al ., 2018) in cereals. A highly efficient approach combining PacBio Iso‐Seq of the wild‐type parental genotype with RNA sequencing of multiple mutants was used to clone the wheat leaf rust R gene Lr9 (Wang et al ., 2023). This approach substantially decreased the cost and time of gene cloning, mitigating the necessity to sort chromosomes and is highly amenable for non‐reference wheat R gene donor lines.…”

“…It would be highly interesting to identify the amino acids that determine pathogen specificity and see whether this would allow engineering of a dual‐pathogen receptor. In a similar vein, the tandem kinases Rwt4 and Pm24 have been recently described to be alleles of the same gene but protecting against wheat blast and powdery mildew, respectively (Arora et al ., 2023; Lu et al ., 2020), whereas the leaf rust and powdery mildew R genes Lr9 (Wang et al ., 2023) and Pm57 (Liu et al ., 2023), encode tandem kinases fused to a von Willebrand factor A domain and share 88% amino acid homology (Liu et al ., 2023). It can be hypothesized that Rwt4/Pm24 and Lr9/Pm57 are part of a three‐component (indirect) guardee system, where the pseudokinase in the tandem‐kinase configuration acts as sensor of pathogen effectors.…”

“…Morex delimited the candidate locus to two possible genes (Dinh et al, 2022). Similarly, cloning and characterization of R genes encoding kinase fusion proteins, such as Resistance to Puccinia graminis 1 (Rpg1) (Brueggeman et al, 2002), Yellow rust resistance 15 (Yr15) (Klymiuk et al, 2015), Lr9 (Wang et al, 2023), Stem rust resistance 43 (Sr43) (Yu et al, 2023) Dinh et al (2022) studies suggest that as many as 35% of major, dominant, racespecific R genes to biotrophic pathogens of wheat and barley do not encode canonical NLR immune receptors (Table S3). Many of these (18 out of 85, 20%) encode protein kinase proteins containing fusion domains.…”

A single NLR gene confers resistance to leaf and stripe rust in wheat

“…Exploitation of the genetic reservoir in the secondary gene pool, such as rye and Aegilops species with genomes C,M,N,S, and U, has been limited owing to the lack of homoeologous recombination and the presence of gametocidal genes that limit interspecies hybridization [11][12][13] . The recent advancements of genomic-based gene isolation approaches [14][15][16][17][18][19][20][21][22] led to a marked increase in cloning of disease-resistance genes, including from species outside the primary wheat gene pool. To date, 32 rust-resistance genes have been cloned, with more than a half in the last 5 years, and many belonging to plants in the secondary wheat gene pools 23 .…”

“…To isolate the gene, we took a mutational transcriptomics approach 19,22 using six EMS-mutated lines (Extended Data Table 2), D42, and the R gene donor Ae. sharonensis AEG-548-4.…”