The wheat Pm3 resistance gene against the powdery mildew pathogen occurs as an allelic series encoding functionally different immune receptors which induce resistance upon recognition of isolate-specific avirulence (AVR) effectors from the pathogen. Here, we describe the identification of five effector proteins from the mildew pathogens of wheat, rye, and the wild grass Dactylis glomerata , specifically recognized by the PM3B, PM3C and PM3D receptors. Together with the earlier identified AVRPM3 A2/F2 , the recognized AVRs of PM3B/C, (AVRPM3 B2/C2 ), and PM3D (AVRPM3 D3 ) belong to a large group of proteins with low sequence homology but predicted structural similarities. AvrPm3 b2/c2 and AvrPm3 d3 are conserved in all tested isolates of wheat and rye mildew, and non-host infection assays demonstrate that Pm3b , Pm3c , and Pm3d are also restricting the growth of rye mildew on wheat. Furthermore, divergent AVR homologues from non-adapted rye and Dactylis mildews are recognized by PM3B, PM3C, or PM3D, demonstrating their involvement in host specificity.

Aegilops tauschii, the diploid wild progenitor of the D subgenome of bread wheat, is a reservoir of genetic diversity for improving bread wheat performance and environmental resilience. Here we sequenced 242 Ae. tauschii accessions and compared them to the wheat D subgenome to characterize genomic diversity. We found that a rare lineage of Ae. tauschii geographically restricted to present-day Georgia contributed to the wheat D subgenome in the independent hybridizations that gave rise to modern bread wheat. Through k-mer-based association mapping, we identified discrete genomic regions with candidate genes for disease and pest resistance and demonstrated their functional transfer into wheat by transgenesis and wide crossing, including the generation of a library of hexaploids incorporating diverse Ae. tauschii genomes. Exploiting the genomic diversity of the Ae. tauschii ancestral diploid genome permits rapid trait discovery and functional genetic validation in a hexaploid background amenable to breeding.

Crop breeding for resistance to pathogens largely relies on genes encoding receptors that confer race-specific immunity. Here we report the identification of the wheat Pm4 racespecific resistance gene to powdery mildew. Pm4 encodes a putative chimeric protein of a serine-threonine kinase and multiple C2-domains and transmembrane regions, a unique domain architecture among known resistance proteins. Pm4 undergoes constitutive alternative splicing generating two isoforms with different protein domain topologies that are both essential for resistance function. Both isoforms interact and localize to the endoplasmatic reticulum (ER) when co-expressed. Pm4 reveals additional diversity of immune receptor architecture to be explored for breeding and suggests an ER-based molecular mechanism of Pm4-mediated race-specific resistance.

Aegilops tauschii, the diploid wild progenitor of the D-subgenome of bread wheat, constitutes a reservoir of genetic diversity for improving bread wheat performance and environmental resilience. To better define and understand this diversity, we sequenced 242 Ae. tauschii accessions and compared them to the wheat D-subgenome. We characterized a rare, geographically-restricted lineage of Ae. tauschii and discovered that it contributed to the wheat D-subgenome, thereby elucidating the origin of bread wheat from at least two independent hybridizations. We then used k-mer-based association mapping to identify discrete genomic regions with candidate genes for disease and pest resistance and demonstrated their functional transfer into wheat by transgenesis and wide crossing, including the generation of a library of ‘synthetic’ hexaploids incorporating diverse Ae. tauschii genomes. This pipeline permits rapid trait discovery in the diploid ancestor through to functional genetic validation in a hexaploid background amenable to breeding.

Plant genomes have evolved several evolutionary mechanisms to tolerate and make use of transposable elements (TEs). Of these, transposon domestication into cis-regulatory and microRNA (miRNA) sequences is proposed to contribute to abiotic/biotic stress adaptation in plants. The wheat genome is derived at 85% from TEs, and contains thousands of miniature inverted-repeat transposable elements (MITEs), whose sequences are particularly prone for domestication into miRNA precursors. In this study, we investigate the contribution of TEs to the wheat small RNA immune response to the lineage-specific, obligate powdery mildew pathogen. We show that MITEs of the Mariner superfamily contribute the largest diversity of miRNAs to the wheat immune response. In particular, MITE precursors of miRNAs are wide-spread over the wheat genome, and highly conserved copies are found in the Lr34 and QPm.tut-4A mildew resistance loci. Our work suggests that transposon domestication is an important evolutionary force driving miRNA functional innovation in wheat immunity.