| | Plants respond to pathogen attack with complex signaling and defense mechanisms including hypersensitive response, which results in rapid cell death. Numerous mutants, resulting in unregulated cell death, have been identified in many species. In barley, such mutants are called "necrotic" while in Arabidopsis they are labeled "lesion mimic" to suggest their involvement in mimicking response to pathogen attack. Lesion mimic or necrotic mutants have been extensively characterized in maize (reviewed in Johal et al., Bioessays 17:685, 1995) and Arabidopsis (reviewed in Lorrain et al., Trends Plant Sci. 8:263, 2003). In barley, the most famous necrotic mutant is mlo (Wolter et al., Mol. Gen. Genet. 239:122, 1993), but otherwise such mutants have received only limited attention. The wild-type Mlo gene encodes a unique membrane anchored protein with six membrane-spanning helices and a postulated dual negative control function in leaf cell death and onset of pathogen defense (Buschges et al., Cell 88:695, 1997). The recessive mlo allele confers durable broad-spectrum resistance to almost all known isolates of the biotrophic fungal pathogen Erysiphe graminis f. sp. hordei (powdery mildew) (Jorgensen, Euphytica 63:141, 1992). However, all mlo lines are hyper-susceptible to the hemibiotrophic fungi Bipolaris sorokiniana (teleomorph Cochliobolus sativus) and Magnaporthe grisea (Kumar et al., Phytopath. 91:127, 2001; Jarosch et al., Mol. Plant-Microbe Interact. 12:508, 1999). Since the survival of hemibiotrophic pathogens in their necrotrophic phase depends on host cell death, the lack of Mlo gene function may antagonize plant defenses to these organisms. The simplest interpretation of these observations may be that increased susceptibility to cell death, as in necrotic mutants, may be antagonistic to biotrophic organisms and favor necrotrophic organisms. Here we report the identification of fast neutron induced mutants FN085 and FN338 as allelic to the barley nec1 locus. By homology to the Arabidopsis Hlm1 gene, the Nec1 gene encodes the cyclic nucleotide-gated ion channel (CNGC) 4 protein (Rostoks et al., submitted). This protein belongs to a family of proteins that are weakly selective cation channels, permeable to K+, Na+ and/or Ca++ and regulated by cyclic nucleotides and calmodulin. In Arabidopsis the hlm1 allele confers increased resistance to Pseudomonas syringae pv. tomato (Balague et al., Plant Cell 12:365, 2003). We tested these mutants and several other necrotic mutants for their reaction to stem rust Puccinia graminis f. sp. tritici (B. Steffenson, unpublished). The CNGC4 mutants FN085, in susceptible cv. Steptoe background, and FN338, in resistant cv. Morex background, did not differ from the wild-type in their reaction to the stem rust pathotype MCC, thus this mutation did not affect susceptibility or resistance to the pathogen. Surprisingly four other fast neutron-induced necrotic mutants, all in susceptible cv. Steptoe background, showed remarkable resistance to the stem rust pathotype MCC, while several others showed no change in their response when compared to the wild-type. In order to identify the genes involved, the four resistant mutants were subjected to analysis on the Barley 1 Affymetrix microarray. Preliminary data indicate that several genes are deleted in each mutant. These could be multiple deletions at several loci or due to one large deletion. To simplify the analysis, the mutants were backcrossed to wild-type and reselected for new analysis on the microarray. The mutants are also being tested for response to different pathogens.
A. Kleinhofs (1), N. Rostoks (2), L. Zhang (1) and B. Steffenson (3)
(1) Dept. Crop and Soil Sciences and School of Molecular Biosciences, Washington State University, Pullman, WA 99164-6420, USA
(2) Scottish Crop Research Institute, Invergowrie, Dundee, DD2 5DA, United Kingdom
(3) Department of Plant Pathology, University of Minnesota, St. Paul, MN 55108-6030, USA
Presented at the 18th North American Barley Researchers Workshop, July 17-20, 2005 |
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