Liselotte L. Selter

A Putative ABC Transporter Confers Durable Resistance to Multiple Fungal Pathogens in Wheat

Agricultural crops benefit from resistance to pathogens that endures over years and generations of both pest and crop. Durable disease resistance, which may be partial or complete, can be controlled by several genes. Some of the most devastating fungal pathogens in wheat are leaf rust, stripe rust, and powdery mildew. The wheat gene Lr34 has supported resistance to these pathogens for more than 50 years. Lr34 is now shared by wheat cultivars around the world. Here, we show that the LR34 protein resembles adenosine triphosphate–binding cassette transporters of the pleiotropic drug resistance subfamily. Alleles of Lr34 conferring resistance or susceptibility differ by three genetic polymorphisms. The Lr34 gene, which functions in the adult plant, stimulates senescence-like processes in the flag leaf tips and edges.

Lr34 multi-pathogen resistance ABC transporter: molecular analysis of homoeologous and orthologous genes in hexaploid wheat and other grass species

The Triticum aestivum (bread wheat) disease resistance gene Lr34 confers durable, race non-specific protection against three fungal pathogens, and has been a highly relevant gene for wheat breeding since the green revolution. Lr34, located on chromosome 7D, encodes an ATP-binding cassette (ABC) transporter. Both wheat cultivars with and without Lr34-based resistance encode a putatively functional protein that differ by only two amino acid polymorphisms. In this study, we focused on the identification and characterization of homoeologous and orthologous Lr34 genes in hexaploid wheat and other grasses. In hexaploid wheat we found an expressed and putatively functional Lr34 homoeolog located on chromosome 4A, designated Lr34-B. Another homoeologous Lr34 copy, located on chromosome 7A, was disrupted by the insertion of repetitive elements. Protein sequences of LR34-B and LR34 were 97% identical. Orthologous Lr34 genes were detected in the genomes of Oryza sativa (rice) and Sorghum bicolor (sorghum). Zea mays (maize), Brachypodium distachyon and Hordeum vulgare (barley) lacked Lr34 orthologs, indicating independent deletion of this particular ABC transporter. Lr34 was part of a gene-rich island on the wheat D genome. We found gene colinearity on the homoeologous A and B genomes of hexaploid wheat, but little microcolinearity in other grasses. The homoeologous LR34-B protein and the orthologs from rice and sorghum have the susceptible haplotype for the two critical polymorphisms distinguishing the LR34 proteins from susceptible and resistant wheat cultivars. We conclude that the particular Lr34-haplotype found in resistant wheat cultivars is unique. It probably resulted from functional gene diversification that occurred after the polyploidization event that was at the origin of cultivated bread wheat.

Functional variability of the Lr34 durable resistance gene in transgenic wheat.

Breeding for durable disease resistance is challenging, yet essential to improve crops for sustainable agriculture. The wheat Lr34 gene is one of the few cloned, durable resistance genes in plants. It encodes an ATP binding cassette transporter and has been a source of resistance against biotrophic pathogens, such as leaf rust (Puccinina triticina), for over 100 years. As endogenous Lr34 confers quantitative resistance, we wanted to determine the effects of transgenic Lr34 with specific reference to how expression levels affect resistance. Transgenic Lr34 wheat lines were made in two different, susceptible genetic backgrounds. We found that the introduction of the Lr34 resistance allele was sufficient to provide comparable levels of leaf rust resistance as the endogenous Lr34 gene. As with the endogenous gene, we observed resistance in seedlings after cold treatment and in flag leaves of adult plants, as well as Lr34-associated leaf tip necrosis. The transgene-based Lr34 resistance did not involve a hypersensitive response, altered callose deposition or up-regulation of PR genes. Higher expression levels compared to endogenous Lr34 were observed in the transgenic lines both at seedling as well as adult stage and some improvement of resistance was seen in the flag leaf. Interestingly, in one genetic background the transgenic Lr34-based resistance resulted in improved seedling resistance without cold treatment. These data indicate that functional variability in Lr34-based resistance can be created using a transgenic approach.

The wheat resistance gene Lr34 results in the constitutive induction of multiple defense pathways in transgenic barley

The wheat gene Lr34 encodes an ABCG-type transporter which provides durable resistance against multiple pathogens. Lr34 is functional as a transgene in barley, but its mode of action has remained largely unknown both in wheat and barley. Here we studied gene expression in uninfected barley lines transgenic for Lr34. Genes from multiple defense pathways contributing to basal and inducible disease resistance were constitutively active in seedlings and mature leaves. In addition, the hormones jasmonic acid and salicylic acid were induced to high levels, and increased levels of lignin as well as hordatines were observed. These results demonstrate a strong, constitutive re-programming of metabolism by Lr34. The resistant Lr34 allele (Lr34res) encodes a protein that differs by two amino acid polymorphisms from the susceptible Lr34sus allele. The deletion of a single phenylalanine residue in Lr34sus was sufficient to induce the characteristic Lr34-based responses. Combination of Lr34res and Lr34sus in the same plant resulted in a reduction of Lr34res expression by 8- to 20-fold when the low-expressing Lr34res line BG8 was used as a parent. Crosses with the high-expressing Lr34res line BG9 resulted in an increase of Lr34sus expression by 13- to 16-fold in progenies that inherited both alleles. These results indicate an interaction of the two Lr34 alleles on the transcriptional level. Reduction of Lr34res expression in BG8 crosses reduced the negative pleiotropic effects of Lr34res on barley growth and vigor without compromising disease resistance, suggesting that transgenic combination of Lr34res and Lr34sus can result in agronomically useful resistance.

Genomic Approaches Towards Durable Fungal Disease Resistance in Wheat

In the last years there has been enormous progress in the molecular understanding of fungal disease resistance in plants. Research on effector-based immunity which is mediated by major resistance (R) genes has been greatly stimulated by the molecular isolation of plant resistance genes as well as the first fungal effectors. In addition, the first genes underlying QTLs or partial disease resistance have been cloned. However, much of this work is still in a phase of basic research and there is a need for translational approaches to realize the globally needed improvements of disease resistance in wheat. In particular, it is essential that future strategies are aiming at achieving durable resistance against pathogens. Durable resistance has been defined by Johnson (Genetic background of durable resistance. In: Lamberti F, Waller JM, Van der Graaff NA (eds) Durable resistance in crops. Plenum, New York, pp 5–24, 1983) as a resistance which remains effective in cultivars that are widely grown for long periods and in environments favorable to the disease. In this article we will discuss different molecular strategies towards achieving durable disease resistance in wheat. In particular, our group focuses on the Pm3 allelic series of race-specific powdery mildew R genes and the Lr34/Yr18/Pm38/Sr57 race non-specific multi-pathogen resistance gene.

Identification and Implementation of Resistance: Genomics-Assisted use of Genetic Resources for Breeding Against Powdery Mildew and Stagonospora Nodorum Blotch in Wheat

Wheat belongs to the three most important cereal crops of the world and is grown under a wide variety of climatic and agricultural conditions. Fungal pathogens represent the most relevant biotic stresses for wheat. These include different rust species, powdery mildew, leaf spots, as well as a number of other diseases that result in reduced grain yield and quality. Recently developed genomic tools allow new approaches to improve breeding for resistance to these pathogens based on a more efficient use of genetic resources. In this chapter, we will focus on the powdery mildew and Stagonospora nodorum blotch diseases and discuss the successful identification of wheat genes determining the outcome of pathogen-host interaction and the development of perfect markers for them. Genomic approaches, including gene cloning, allele mining, transcriptomics and comparative genomics have greatly changed and improved our understanding of molecular wheat-powdery mildew interactions. For the necrotrophic pathogen Stagonospora nodorum much of the interaction was found to be based on pathogen toxins and host susceptibility genes. The work on specific gene-for-gene interactions opened new possibilities for more efficient resistance breeding. In addition, the molecular identification of quantitatively acting resistance loci in wheat has made important progress, although only few such genes have been cloned, only one of them each against mildew and Stagonospora nodorum blotch. However, even at this early stage it can be foreseen that the new knowledge might revolutionize breeding for durable resistance in the near future. The progress made towards a whole genome sequence of wheat together with ongoing developments of high throughput techniques provides a completely new perspective on resistance breeding against these two diseases.