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A compatible interaction of Alternaria brassicicola with Arabidopsis thaliana ecotype DiG: evidence for a specific transcriptional signature.

Mukherjee AK, Lev S, Gepstein S, Horwitz BA - BMC Plant Biol. (2009)

Bottom Line: PR1, and a monooxygenase gene identified in this study, MO1, are preferentially up-regulated in the compatible interaction.In contrast, GLIP1, which encodes a secreted lipase, and DIOX1, a pathogen-response related dioxygenase, are preferentially up-regulated in the incompatible interaction.The results show that DiG is not only more susceptible, but demonstrate that its interaction with A. brassicicola has a specific transcriptional signature.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Biology, Israel Institute of Technology, Technion, Haifa, Israel. titirtua@gmail.com

ABSTRACT

Background: The interaction of Arabidopsis with Alternaria brassicicola provides a model for disease caused by necrotrophs, but a drawback has been the lack of a compatible pathosystem. Infection of most ecotypes, including the widely-studied line Col-0, with this pathogen generally leads to a lesion that does not expand beyond the inoculated area. This study examines an ecotype, Dijon G (DiG), which is considered sensitive to A. brassicicola.

Results: We show that the interaction has the characteristics of a compatible one, with expanding rather than limited lesions. To ask whether DiG is merely more sensitive to the pathogen or, rather, interacts in distinct manner, we identified genes whose regulation differs between Col-0 and DiG challenged with A. brassicicola. Suppression subtractive hybridization was used to identify differentially expressed genes, and their expression was verified using semi-quantitative PCR. We also tested a set of known defense-related genes for differential regulation in the two plant-pathogen interactions. Several known pathogenesis-related (PR) genes are up-regulated in both interactions. PR1, and a monooxygenase gene identified in this study, MO1, are preferentially up-regulated in the compatible interaction. In contrast, GLIP1, which encodes a secreted lipase, and DIOX1, a pathogen-response related dioxygenase, are preferentially up-regulated in the incompatible interaction.

Conclusion: The results show that DiG is not only more susceptible, but demonstrate that its interaction with A. brassicicola has a specific transcriptional signature.

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Related in: MedlinePlus

Characterization of Arabidopsis-Alternaria brassicicola pairs. a) Symptoms in different ecotypes and genotypes, 3 days after inoculation of intact leaves. Top row, inoculated; bottom row, control. glip1-1 and glip1-2 are two mutants at the glip1 locus encoding a secreted lipase [28]; acd1 is a lesion mimic mutant [32]. Scale bar indicates 2 cm. b) Magnification of images of leaves from (a) showing the ring-like pattern in the progression of the lesion on a DiG leaf (arrows). The innermost dark, thin, arc (no arrow) is material from the inoculum. Scale bar indicates 1 cm. c) Size of lesions on Col-0 and DiG leaves at different times post-inoculation. Representative infected leaves are shown, photographed at the indicated times after inoculation. Scale bar indicates 2 cm. d) Quantitative analysis of lesion size and spore production. Top panel, lesion diameter was measured 5 days after inoculation. Error bars indicate standard errors of the mean for 7 replicate lesions (for Col-0, 9 and DiG, 10 replicates). Lower panel, lesions were excised 5 days after inoculation, the conidia suspended in water, and counted under the microscope in a hemocytometer chamber. Values are means of two independent experiments, consisting of 12 and 4–5 replicates, respectively; the error bars indicate the standard error of the mean of the combined data from the two experiments.
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Figure 1: Characterization of Arabidopsis-Alternaria brassicicola pairs. a) Symptoms in different ecotypes and genotypes, 3 days after inoculation of intact leaves. Top row, inoculated; bottom row, control. glip1-1 and glip1-2 are two mutants at the glip1 locus encoding a secreted lipase [28]; acd1 is a lesion mimic mutant [32]. Scale bar indicates 2 cm. b) Magnification of images of leaves from (a) showing the ring-like pattern in the progression of the lesion on a DiG leaf (arrows). The innermost dark, thin, arc (no arrow) is material from the inoculum. Scale bar indicates 1 cm. c) Size of lesions on Col-0 and DiG leaves at different times post-inoculation. Representative infected leaves are shown, photographed at the indicated times after inoculation. Scale bar indicates 2 cm. d) Quantitative analysis of lesion size and spore production. Top panel, lesion diameter was measured 5 days after inoculation. Error bars indicate standard errors of the mean for 7 replicate lesions (for Col-0, 9 and DiG, 10 replicates). Lower panel, lesions were excised 5 days after inoculation, the conidia suspended in water, and counted under the microscope in a hemocytometer chamber. Values are means of two independent experiments, consisting of 12 and 4–5 replicates, respectively; the error bars indicate the standard error of the mean of the combined data from the two experiments.

Mentions: A total of five wild type ecotypes, Col-0, Col-6, DiG (Dijon G), Ler (Landsberg erecta), Ws (Wassilewskija) and three mutants (glip1-1, glip1-2 and acd1) were screened against A. brassicicola (Fig. 1a). These mutants were tested initially, because glip1 mutants were shown to be susceptible to A. brassicicola [28]. The mutant acd1 was chosen because we reasoned that, as LLS1 in maize and its ortholog ACD1 in Arabidopsis are required to limit the spread of cell death [31,32], loss of this gene might increase the spread of a necrotrophic pathogen. Inoculum amount and sampling times after inoculation were calibrated in preliminary experiments so that the different plant-fungal pairs could be compared under non-saturating conditions (data not shown). Lesion diameter and spore production were measured (Fig. 1b) to assess the disease progression. The Col-0 accession showed an incompatible interaction, in which the lesion did not progress beyond the boundaries of the inoculated region. Accession DiG was most susceptible, showing larger lesions than either of the glip1 mutants or Col-6, which are relatively susceptible as compared to Col-0 [28]. The lesion-mimic mutant acd1, was no more susceptible to the pathogen than was Col-0 (Fig. 1a, d). The lesions on DiG leaves continued to spread (Fig. 1c) and often show concentric rings (Fig. 1b), as seen in the interaction with the compatible host Brassica oleracea but not in incompatible interactions with Arabidopsis. To test the possibility that the DiG-A. brassicicola interaction has a unique transcriptional signature, two approaches were followed: differential library screening and a candidate gene approach.


A compatible interaction of Alternaria brassicicola with Arabidopsis thaliana ecotype DiG: evidence for a specific transcriptional signature.

Mukherjee AK, Lev S, Gepstein S, Horwitz BA - BMC Plant Biol. (2009)

Characterization of Arabidopsis-Alternaria brassicicola pairs. a) Symptoms in different ecotypes and genotypes, 3 days after inoculation of intact leaves. Top row, inoculated; bottom row, control. glip1-1 and glip1-2 are two mutants at the glip1 locus encoding a secreted lipase [28]; acd1 is a lesion mimic mutant [32]. Scale bar indicates 2 cm. b) Magnification of images of leaves from (a) showing the ring-like pattern in the progression of the lesion on a DiG leaf (arrows). The innermost dark, thin, arc (no arrow) is material from the inoculum. Scale bar indicates 1 cm. c) Size of lesions on Col-0 and DiG leaves at different times post-inoculation. Representative infected leaves are shown, photographed at the indicated times after inoculation. Scale bar indicates 2 cm. d) Quantitative analysis of lesion size and spore production. Top panel, lesion diameter was measured 5 days after inoculation. Error bars indicate standard errors of the mean for 7 replicate lesions (for Col-0, 9 and DiG, 10 replicates). Lower panel, lesions were excised 5 days after inoculation, the conidia suspended in water, and counted under the microscope in a hemocytometer chamber. Values are means of two independent experiments, consisting of 12 and 4–5 replicates, respectively; the error bars indicate the standard error of the mean of the combined data from the two experiments.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC2664814&req=5

Figure 1: Characterization of Arabidopsis-Alternaria brassicicola pairs. a) Symptoms in different ecotypes and genotypes, 3 days after inoculation of intact leaves. Top row, inoculated; bottom row, control. glip1-1 and glip1-2 are two mutants at the glip1 locus encoding a secreted lipase [28]; acd1 is a lesion mimic mutant [32]. Scale bar indicates 2 cm. b) Magnification of images of leaves from (a) showing the ring-like pattern in the progression of the lesion on a DiG leaf (arrows). The innermost dark, thin, arc (no arrow) is material from the inoculum. Scale bar indicates 1 cm. c) Size of lesions on Col-0 and DiG leaves at different times post-inoculation. Representative infected leaves are shown, photographed at the indicated times after inoculation. Scale bar indicates 2 cm. d) Quantitative analysis of lesion size and spore production. Top panel, lesion diameter was measured 5 days after inoculation. Error bars indicate standard errors of the mean for 7 replicate lesions (for Col-0, 9 and DiG, 10 replicates). Lower panel, lesions were excised 5 days after inoculation, the conidia suspended in water, and counted under the microscope in a hemocytometer chamber. Values are means of two independent experiments, consisting of 12 and 4–5 replicates, respectively; the error bars indicate the standard error of the mean of the combined data from the two experiments.
Mentions: A total of five wild type ecotypes, Col-0, Col-6, DiG (Dijon G), Ler (Landsberg erecta), Ws (Wassilewskija) and three mutants (glip1-1, glip1-2 and acd1) were screened against A. brassicicola (Fig. 1a). These mutants were tested initially, because glip1 mutants were shown to be susceptible to A. brassicicola [28]. The mutant acd1 was chosen because we reasoned that, as LLS1 in maize and its ortholog ACD1 in Arabidopsis are required to limit the spread of cell death [31,32], loss of this gene might increase the spread of a necrotrophic pathogen. Inoculum amount and sampling times after inoculation were calibrated in preliminary experiments so that the different plant-fungal pairs could be compared under non-saturating conditions (data not shown). Lesion diameter and spore production were measured (Fig. 1b) to assess the disease progression. The Col-0 accession showed an incompatible interaction, in which the lesion did not progress beyond the boundaries of the inoculated region. Accession DiG was most susceptible, showing larger lesions than either of the glip1 mutants or Col-6, which are relatively susceptible as compared to Col-0 [28]. The lesion-mimic mutant acd1, was no more susceptible to the pathogen than was Col-0 (Fig. 1a, d). The lesions on DiG leaves continued to spread (Fig. 1c) and often show concentric rings (Fig. 1b), as seen in the interaction with the compatible host Brassica oleracea but not in incompatible interactions with Arabidopsis. To test the possibility that the DiG-A. brassicicola interaction has a unique transcriptional signature, two approaches were followed: differential library screening and a candidate gene approach.

Bottom Line: PR1, and a monooxygenase gene identified in this study, MO1, are preferentially up-regulated in the compatible interaction.In contrast, GLIP1, which encodes a secreted lipase, and DIOX1, a pathogen-response related dioxygenase, are preferentially up-regulated in the incompatible interaction.The results show that DiG is not only more susceptible, but demonstrate that its interaction with A. brassicicola has a specific transcriptional signature.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Biology, Israel Institute of Technology, Technion, Haifa, Israel. titirtua@gmail.com

ABSTRACT

Background: The interaction of Arabidopsis with Alternaria brassicicola provides a model for disease caused by necrotrophs, but a drawback has been the lack of a compatible pathosystem. Infection of most ecotypes, including the widely-studied line Col-0, with this pathogen generally leads to a lesion that does not expand beyond the inoculated area. This study examines an ecotype, Dijon G (DiG), which is considered sensitive to A. brassicicola.

Results: We show that the interaction has the characteristics of a compatible one, with expanding rather than limited lesions. To ask whether DiG is merely more sensitive to the pathogen or, rather, interacts in distinct manner, we identified genes whose regulation differs between Col-0 and DiG challenged with A. brassicicola. Suppression subtractive hybridization was used to identify differentially expressed genes, and their expression was verified using semi-quantitative PCR. We also tested a set of known defense-related genes for differential regulation in the two plant-pathogen interactions. Several known pathogenesis-related (PR) genes are up-regulated in both interactions. PR1, and a monooxygenase gene identified in this study, MO1, are preferentially up-regulated in the compatible interaction. In contrast, GLIP1, which encodes a secreted lipase, and DIOX1, a pathogen-response related dioxygenase, are preferentially up-regulated in the incompatible interaction.

Conclusion: The results show that DiG is not only more susceptible, but demonstrate that its interaction with A. brassicicola has a specific transcriptional signature.

Show MeSH
Related in: MedlinePlus