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The F-box protein MAX2 contributes to resistance to bacterial phytopathogens in Arabidopsis thaliana.

Piisilä M, Keceli MA, Brader G, Jakobson L, Jõesaar I, Sipari N, Kollist H, Palva ET, Kariola T - BMC Plant Biol. (2015)

Bottom Line: Interestingly, max2 mutant plants showed increased susceptibility to the bacterial necrotroph Pectobacterium carotovorum as well as to the hemi-biotroph Pseudomonas syringae but not to the fungal necrotroph Botrytis cinerea. max2 mutant phenotype was associated with constitutively increased stomatal conductance and decreased tolerance to apoplastic ROS but also with alterations in hormonal balance.We conclude that the increased susceptibility to P. syringae and P. carotovorum is due to increased stomatal conductance in max2 mutants promoting pathogen entry into the plant apoplast.Additional factors contributing to pathogen susceptibility in max2 plants include decreased tolerance to pathogen-triggered apoplastic ROS and alterations in hormonal signaling.

View Article: PubMed Central - PubMed

ABSTRACT

Background: The Arabidopsis thaliana F-box protein MORE AXILLARY GROWTH2 (MAX2) has previously been characterized for its role in plant development. MAX2 appears essential for the perception of the newly characterized phytohormone strigolactone, a negative regulator of polar auxin transport in Arabidopsis.

Results: A reverse genetic screen for F-box protein mutants altered in their stress responses identified MAX2 as a component of plant defense. Here we show that MAX2 contributes to plant resistance against pathogenic bacteria. Interestingly, max2 mutant plants showed increased susceptibility to the bacterial necrotroph Pectobacterium carotovorum as well as to the hemi-biotroph Pseudomonas syringae but not to the fungal necrotroph Botrytis cinerea. max2 mutant phenotype was associated with constitutively increased stomatal conductance and decreased tolerance to apoplastic ROS but also with alterations in hormonal balance.

Conclusions: Our results suggest that MAX2 previously characterized for its role in regulation of polar auxin transport in Arabidopsis, and thus plant development also significantly influences plant disease resistance. We conclude that the increased susceptibility to P. syringae and P. carotovorum is due to increased stomatal conductance in max2 mutants promoting pathogen entry into the plant apoplast. Additional factors contributing to pathogen susceptibility in max2 plants include decreased tolerance to pathogen-triggered apoplastic ROS and alterations in hormonal signaling.

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Pathogen-triggered stomatal closure is impaired in max2 mutant lines. Four-week old wild-type Col-0 and max2 lines were inoculated with Pseudomonas syringae pv. tomato DC3000. A) Measurement of stomatal aperture of wild-type Col-0 and max2 lines in response to P. syringae. Leaves were first stained with 20 μM propidium iodide (PI) solution and then inoculated with 300 μl of bacterial solution (108 cfu/ml). Stomatal aperture width was measured after indicated time points using ImageJ image processing program. B) Representative pictures of stomatal response of Col-0 and max2 lines under florescent microscope using 20x objective 0, 1, 2 and 4 h after inoculation with the bacteria. Results are shown as the mean (n = 80-100) ± SE. **P < 0.01; two-tailed t test. The experiments were repeated three times with similar results.
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Fig6: Pathogen-triggered stomatal closure is impaired in max2 mutant lines. Four-week old wild-type Col-0 and max2 lines were inoculated with Pseudomonas syringae pv. tomato DC3000. A) Measurement of stomatal aperture of wild-type Col-0 and max2 lines in response to P. syringae. Leaves were first stained with 20 μM propidium iodide (PI) solution and then inoculated with 300 μl of bacterial solution (108 cfu/ml). Stomatal aperture width was measured after indicated time points using ImageJ image processing program. B) Representative pictures of stomatal response of Col-0 and max2 lines under florescent microscope using 20x objective 0, 1, 2 and 4 h after inoculation with the bacteria. Results are shown as the mean (n = 80-100) ± SE. **P < 0.01; two-tailed t test. The experiments were repeated three times with similar results.

Mentions: Stomatal closure in response to invading bacteria such as P. syringae is a well-described component of the innate immunity response in Arabidopsis [6]. The more open stomatal aperture in the absence of stress (Figure 3A) and the enhanced susceptibility of max2 plants to spray-inoculated P. syringae (Figure 5A and B) indicated that the pathogen-triggered stomatal closure could be impaired in these plants. To elucidate this, we infected max2 and wild-type plants with P. syringae bacterial suspension and checked stomatal response to living bacterial cells 0, 1, 2 and 4 h after inoculation using fluorescence microscopy using the method introduced by Chitrakar and Melotto 2010 [54]. When max2 and wild-type leaves were incubated with P. syringae stomatal closure was triggered in wild-type plants1h after infection but this was not observed in max2 lines where the stomatal opening was rather getting higher during measured time points (Figure 6). P. syringae DC3000 has been shown to induce re-opening of the stomata from 3 to 4 h after the initial closure by secreting the phytotoxin coronatine [6]. While this was observed for the wild-type at 4 h time point, in max2 plants the stomatal aperture was even larger 2 and 4 h after the infection (Figure 6). Treatment of max2 and wild-type leaves with MgCl2 buffer solution did not alter the stomatal aperture, but yet, the stomata of max2 plants were clearly more open compared to wild-type (Additional file 1: Figure S5). These results clearly indicate that max2 plants have impaired stomatal closure in response to P. syringae allowing increased numbers of bacteria to enter plant apoplast (Figure 5B) leading to more severe susceptibility.Figure 6


The F-box protein MAX2 contributes to resistance to bacterial phytopathogens in Arabidopsis thaliana.

Piisilä M, Keceli MA, Brader G, Jakobson L, Jõesaar I, Sipari N, Kollist H, Palva ET, Kariola T - BMC Plant Biol. (2015)

Pathogen-triggered stomatal closure is impaired in max2 mutant lines. Four-week old wild-type Col-0 and max2 lines were inoculated with Pseudomonas syringae pv. tomato DC3000. A) Measurement of stomatal aperture of wild-type Col-0 and max2 lines in response to P. syringae. Leaves were first stained with 20 μM propidium iodide (PI) solution and then inoculated with 300 μl of bacterial solution (108 cfu/ml). Stomatal aperture width was measured after indicated time points using ImageJ image processing program. B) Representative pictures of stomatal response of Col-0 and max2 lines under florescent microscope using 20x objective 0, 1, 2 and 4 h after inoculation with the bacteria. Results are shown as the mean (n = 80-100) ± SE. **P < 0.01; two-tailed t test. The experiments were repeated three times with similar results.
© Copyright Policy - open-access
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4340836&req=5

Fig6: Pathogen-triggered stomatal closure is impaired in max2 mutant lines. Four-week old wild-type Col-0 and max2 lines were inoculated with Pseudomonas syringae pv. tomato DC3000. A) Measurement of stomatal aperture of wild-type Col-0 and max2 lines in response to P. syringae. Leaves were first stained with 20 μM propidium iodide (PI) solution and then inoculated with 300 μl of bacterial solution (108 cfu/ml). Stomatal aperture width was measured after indicated time points using ImageJ image processing program. B) Representative pictures of stomatal response of Col-0 and max2 lines under florescent microscope using 20x objective 0, 1, 2 and 4 h after inoculation with the bacteria. Results are shown as the mean (n = 80-100) ± SE. **P < 0.01; two-tailed t test. The experiments were repeated three times with similar results.
Mentions: Stomatal closure in response to invading bacteria such as P. syringae is a well-described component of the innate immunity response in Arabidopsis [6]. The more open stomatal aperture in the absence of stress (Figure 3A) and the enhanced susceptibility of max2 plants to spray-inoculated P. syringae (Figure 5A and B) indicated that the pathogen-triggered stomatal closure could be impaired in these plants. To elucidate this, we infected max2 and wild-type plants with P. syringae bacterial suspension and checked stomatal response to living bacterial cells 0, 1, 2 and 4 h after inoculation using fluorescence microscopy using the method introduced by Chitrakar and Melotto 2010 [54]. When max2 and wild-type leaves were incubated with P. syringae stomatal closure was triggered in wild-type plants1h after infection but this was not observed in max2 lines where the stomatal opening was rather getting higher during measured time points (Figure 6). P. syringae DC3000 has been shown to induce re-opening of the stomata from 3 to 4 h after the initial closure by secreting the phytotoxin coronatine [6]. While this was observed for the wild-type at 4 h time point, in max2 plants the stomatal aperture was even larger 2 and 4 h after the infection (Figure 6). Treatment of max2 and wild-type leaves with MgCl2 buffer solution did not alter the stomatal aperture, but yet, the stomata of max2 plants were clearly more open compared to wild-type (Additional file 1: Figure S5). These results clearly indicate that max2 plants have impaired stomatal closure in response to P. syringae allowing increased numbers of bacteria to enter plant apoplast (Figure 5B) leading to more severe susceptibility.Figure 6

Bottom Line: Interestingly, max2 mutant plants showed increased susceptibility to the bacterial necrotroph Pectobacterium carotovorum as well as to the hemi-biotroph Pseudomonas syringae but not to the fungal necrotroph Botrytis cinerea. max2 mutant phenotype was associated with constitutively increased stomatal conductance and decreased tolerance to apoplastic ROS but also with alterations in hormonal balance.We conclude that the increased susceptibility to P. syringae and P. carotovorum is due to increased stomatal conductance in max2 mutants promoting pathogen entry into the plant apoplast.Additional factors contributing to pathogen susceptibility in max2 plants include decreased tolerance to pathogen-triggered apoplastic ROS and alterations in hormonal signaling.

View Article: PubMed Central - PubMed

ABSTRACT

Background: The Arabidopsis thaliana F-box protein MORE AXILLARY GROWTH2 (MAX2) has previously been characterized for its role in plant development. MAX2 appears essential for the perception of the newly characterized phytohormone strigolactone, a negative regulator of polar auxin transport in Arabidopsis.

Results: A reverse genetic screen for F-box protein mutants altered in their stress responses identified MAX2 as a component of plant defense. Here we show that MAX2 contributes to plant resistance against pathogenic bacteria. Interestingly, max2 mutant plants showed increased susceptibility to the bacterial necrotroph Pectobacterium carotovorum as well as to the hemi-biotroph Pseudomonas syringae but not to the fungal necrotroph Botrytis cinerea. max2 mutant phenotype was associated with constitutively increased stomatal conductance and decreased tolerance to apoplastic ROS but also with alterations in hormonal balance.

Conclusions: Our results suggest that MAX2 previously characterized for its role in regulation of polar auxin transport in Arabidopsis, and thus plant development also significantly influences plant disease resistance. We conclude that the increased susceptibility to P. syringae and P. carotovorum is due to increased stomatal conductance in max2 mutants promoting pathogen entry into the plant apoplast. Additional factors contributing to pathogen susceptibility in max2 plants include decreased tolerance to pathogen-triggered apoplastic ROS and alterations in hormonal signaling.

Show MeSH
Related in: MedlinePlus