Helen G. McFadden

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.

Gene expression profile changes in cotton root and hypocotyl tissues in response to infection with Fusarium oxysporum f. sp. vasinfectum.

Microarray analysis of large-scale temporal and tissue-specific plant gene expression changes occurring during a susceptible plant-pathogen interaction revealed different gene expression profile changes in cotton root and hypocotyl tissues. In hypocotyl tissues infected with Fusarium oxysporum f. sp. vasinfectum, increased expression of defense-related genes was observed, whereas few changes in the expression levels of defense-related genes were found in infected root tissues. In infected roots, more plant genes were repressed than were induced, especially at the earlier stages of infection. Although many known cotton defense responses were identified, including induction of pathogenesis-related genes and gossypol biosynthesis genes, potential new defense responses also were identified, such as the biosynthesis of lignans. Many of the stress-related gene responses were common to both tissues. The repression of drought-responsive proteins such as aquaporins in both roots and hypocotyls represents a previously unreported response of a host to pathogen attack that may be specific to vascular wilt diseases. Gene expression results implicated the phytohormones ethylene and auxin in the disease process. Biochemical analysis of hormone level changes supported this observation.

Leaf tip necrosis, molecular markers and β1-proteasome subunits associated with the slow rusting resistance genes Lr46/Yr29

Resistance based on slow-rusting genes has proven to be a useful strategy to develop wheat cultivars with durable resistance to rust diseases in wheat. However this type of resistance is often difficult to incorporate into a single genetic background due to the polygenic and additive nature of the genes involved. Therefore, markers, both molecular and phenotypic, are useful tools to facilitate the use of this type of resistance in wheat breeding programs. We have used field assays to score for both leaf and yellow rust in an Avocet-YrA × Attila population that segregates for several slow-rusting leaf and yellow rust resistance genes. This population was analyzed with the AFLP technique and the slow-rusting resistance locus Lr46/Yr29 was identified. A common set of AFLP and SSR markers linked to the Lr46/Yr29 locus was identified and validated in other recombinant inbred families developed from single chromosome recombinant populations that segregated for Lr46. These populations segregated for leaf tip necrosis (LTN) in the field, a trait that had previously been associated with Lr34/Yr18. We show that LTN is also pleiotropic or closely linked to the Lr46/Yr29 locus and suggest that a new Ltn gene designation should be given to this locus, in addition to the one that already exists for Lr34/Yr18. Coincidentally, members of a small gene family encoding β-1 proteasome subunits located on group 1L and 7S chromosomes implicated in plant defense were linked to the Lr34/Yr18 and Lr46/Yr29 loci.

Autoactive alleles of the flax L6 rust resistance gene induce non-race-specific rust resistance associated with the hypersensitive response.

L6 is a nucleotide binding site-leucine rich repeat (NBS-LRR) gene that confers race-specific resistance in flax (Linum usitatissimum) to strains of flax rust (Melampsora lini) that carry avirulence alleles of the AvrL567 gene but not to rust strains that carry only the virulence allele. Several mutant and recombinant forms of L6 were made that altered either the methionine-histidine-aspartate (MHD) motif conserved in the NBS domain of resistance proteins or exchanged the short domain C-terminal to the LRR region that is highly variable among L allele products. In transgenic flax some of these alleles are autoactive; they cause a gene dosage-dependent dwarf phenotype and constitutive expression of genes that are markers for the plant defense response. Their effects and penetrance ranged from extreme to mild in their degree of plant stunting, survival, and reproduction. Dwarf plants were also resistant to flax rust strains virulent to wild-type L6 plants, and this nonspecific resistance was associated with a hypersensitive response (HR) at the site of rust infection. The strongest autoactive allele, expressed in Arabidopsis from an ethanol-inducible promoter, gave rise to plant death dependent on the enhanced disease susceptibility 1 (EDS1) gene, which indicates that the mutant flax (Linaceae) L6 gene can signal cell death through a defined disease-resistance pathway in a different plant family (Brassicaceae).

Fusarium wilt (Fusarium oxysporum f. sp. vasinfectum) genes expressed during infection of cotton (Gossypium hirsutum)

SUMMARY We sought to identify Fusarium oxysporum f. sp. vasinfectum (Fov) genes that may be associated with pathogenicity. Initially we utilized microarray and Q-PCR technology to identify Fov genes expressed in root and hypocotyl tissues during a compatible infection of cotton. We identified 218 fungal clones representing 174 Fov non-redundant genes as expressed in planta. The majority of the expressed sequences were expressed in infected roots, with only six genes detected in hypocotyl tissue. The Fov genes identified were predominately of unknown function or associated with fungal growth and energy production. We then analysed the expression of the identified fungal genes in infected roots and in saprophytically grown mycelia and identified 11 genes preferentially expressed in plant tissue. A putative oxidoreductase gene (with homology to AtsC) was found to be highly preferentially expressed in planta. In Agrobacterium tumefaciens, AtsC is associated with virulence. Inoculation of a susceptible and a partially resistant cotton cultivar with either a pathogenic or a non-pathogenic isolate of Fov revealed that the expression of the Fov AtsC homologue was associated with pathogenicity and disease symptom formation.

Assessment of Gossypium sturtianum and G. australe as potential sources of Fusarium wilt resistance to cotton

When challenged with Fusarium oxysporum f. sp. vasinfectum (Fov) from vegetative compatibility groups (VCGs) 01111 and 01112 in glasshouse tests, Gossypium australe Mueller and Gossypium sturtianum Willis accessions showed a variety of disease responses ranging from highly resistant to highly susceptible. Under high disease pressure G. sturtianum accession Gos-5275 was significantly more resistant than the commercial G. hirsutum cultivars that are designated standards for Fusarium resistance by Australian cotton breeders. Under low disease pressure G. sturtianum accession Gos-5250 was more susceptible than a highly susceptible commercial cultivar. A series of glasshouse tests was performed at two locations (Indooroopilly, QLD. and Canberra, ACT), and under low and high disease pressure. In these tests, a hexaploid cross (Gos-5271) generated from a Fusarium-resistant G. sturtianum (Gos-5275) and a Fusarium-susceptible G. hirsutum L. (CPI-138969) was significantly more resistant to Fusarium wilt than its G. hirsutum parent. Thus G. sturtianum, with a diploid genome and a range of responses to Fov challenge, has the potential to provide the basis for the elucidation of the genetic basis of resistance to Fusarium wilt in cotton species. In addition, resistant accessions of G. sturtianum are identified as a potential source of Fusarium wilt resistance genes for cotton breeding. In the glasshouse tests used to assess the resistance of various Gossypium accessions to Fusarium wilt disease, the scoring of vascular browning was found to give a more reliable indication of disease severity than the scoring of foliar symptoms.

Wheat rust resistance research at CSIRO

Although chemical control is available for rust diseases in wheat, economic and environmental factors favour genetic solutions. Maintenance and improvement of levels of resistance and durability of the genetic control of the 3 wheat rust diseases will occur with the application of DNA markers for pyramiding resistance genes. Information about the molecular basis of rust resistance, including durable, adult-plant resistance, coming from studies in model species such as flax and flax rust and from studies of wheat and barley, will provide knowledge for new biotechnological approaches to rust resistance. Increasing cereal gene sequence data will improve the efficiency of cloning disease resistance genes and, together with the rapid progress in understanding the molecular basis of rust resistance, will make it possible to construct transgenic plants with multiple rust resistance genes at a single locus, which will provide efficient breeding and increased durability of rust resistance.

The effect of Gossypium C-genome chromosomes on resistance to fusarium wilt in allotetraploid cotton

Fusarium oxysporum f. sp. vasinfectum (Fov) has the potential to become the most economically significant pathogen of cotton in Australia. Although the levels of resistance present in the new commercial cultivars have improved significantly, they are still not immune and cotton breeders continue to look for additional sources of resistance. The native Australian Gossypium species represent an alternative source of resistance because they could have co-evolved with the indigenous Fov pathogens. Forty-six BC3 G. hirsutum × G. sturtianum multiple alien-chromosome-addition-line (MACAL) families were challenged with a field-derived Fov isolate (VCG-01111). The G. hirsutum parent of the hexaploid MACAL is highly susceptible to fusarium wilt; the G. sturtianum parent is strongly resistant. Twenty-two of the BC3 families showed enhanced fusarium wilt resistance relative to the susceptible G. hirsutum parent. Logistic regression identified four G. sturtianum linkage groups with a significant effect on fusarium wilt resistance: two linkage groups were associated with improved fusarium wilt resistance, while two linkage groups were associated with increased fusarium wilt susceptibility.

Cyanoacrylate inhibitors as probes for the nature of the photosystem II herbicide binding site

Three series of phenyl-substituted 2-cyanoacrylates were evaluated using simple quantitative structure activity relationships (QSAR) in an attempt to elucidate the nature of the regions of the binding site occupied by different parts of the molecules. Inhibition of the Hill reaction by substituted 3-phenylamino-2-cyanoacrylates correlated well with the lipophilicity of the substituent. The hydrophobic effect was also dominant when the Hill activity of a series of 3-benzylamino-2-cyanoacrylates was analyzed, although potency was considerably higher in the latter series. Lipophilicity and the electronic nature of the substituents were not major determinants in the Hill inhibitory activity of a series of substituted phcnoxycthyl 2-cyanoacrylic esters. In this case, a significant correlation was found with the molar rcfractivity (MR) of meta substituents, a parameter reflecting substituent size. The results indicate that the phenyl moiety of substituted 3-phenylamino- and 3-bcnzyl- amino-2-cyanoacrylates interacts with an essentially lipophilic binding domain, though it is likely that the two series are oriented differently with the 3-bcnzylamino series able to bind with greater affinity. In the phcnoxycthyl ester series, the substituted phenyl group interacts with a different environment, wherein ortho- and we7tf-substitution is tolerated, dependent on the bulk of the substituent, but /wra-substitution is detrimental to affinity for this region of the site.

Differential induction of chitinase activity in flax (Linum usitatissimum) in response to inoculation with virulent or avirulent strains of Melampsora lini, the cause of flax rust

Chitinase enzymes are induced in many plants in response to pathogen challenge and other stress stimuli. Chitinase induction in flax leaves in response to inoculation with flax rust, caused by Melampsora lini (Pers.) Lev., was investigated in a line of flax (Forge) that has four resistance genes, viz. L6, M, N and P2. Four avirulent rust strains, each of which interacts with just one of the resistance genes in Forge, as well as a strain that is virulent on Forge, were used. Thus chitinase levels associated with resistance reactions triggered by the L6, M, N or P2 genes, and by a susceptible reaction, have been compared in the same host genotype. A marked increase in chitinase activity in inoculated leaves was observed with all four resistance reactions, with the increase occurring earlier with the P2 resistance reaction compared with the L6, M, and N reactions. A moderate increase in chitinase activity was also observed in systemic (new-growth) leaves of flax plants inoculated with strains interacting with the M, N or P2 genes. Leaves inoculated with a virulent strain of rust also had increased chitinase activity but the increase was much less than that found in leaves inoculated with the avirulent strains.