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Characterization of a Novel Cotton Subtilase Gene GbSBT1 in Response to Extracellular Stimulations and Its Role in Verticillium Resistance.

Duan X, Zhang Z, Wang J, Zuo K - PLoS ONE (2016)

Bottom Line: Verticillium pathogens secrete various disease-causing effectors in cotton.Moreover, the GbSBT1 protein is mainly localized in the cell membrane and moves into the cytoplasm following jasmonic acid and ethylene treatments.In summary, GbSBT1 recognizes the effector PHB protein secreted from V. dahliae and is involved in Verticillium-induced resistance in cotton.

View Article: PubMed Central - PubMed

Affiliation: Plant Biotechnology Research Center, School of Agriculture and Life Sciences, Shanghai Jiao Tong University, Shanghai, China.

ABSTRACT
Verticillium wilt is a disastrous vascular disease in plants caused by Verticillium dahliae. Verticillium pathogens secrete various disease-causing effectors in cotton. This study identified a subtilase gene GbSBT1 from Gossypium babardense and investigated the roles against V. dahliae infection. GbSBT1 gene expression is responsive to V. dahliae defense signals, jasmonic acid, and ethylene treatments. Moreover, the GbSBT1 protein is mainly localized in the cell membrane and moves into the cytoplasm following jasmonic acid and ethylene treatments. Silencing GbSBT1 gene expression through virus-induced GbSBT1 gene silencing reduced the tolerance of Pima-90 (resistant genotype), but not facilitated the infection process of V. dahliae in Coker-312 (sensitive genotype). Moreover, the ectopically expressed GbSBT1 gene enhanced the resistance of Arabidopsis to Fusarium oxysporum and V. dahliae infection and activated the expression levels of defense-related genes. Furthermore, pull-down, yeast two-hybrid assay, and BiFC analysis revealed that GbSBT1 interacts with a prohibitin (PHB)-like protein expressed in V. dahliae pathogens during infection. In summary, GbSBT1 recognizes the effector PHB protein secreted from V. dahliae and is involved in Verticillium-induced resistance in cotton.

No MeSH data available.


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Enhanced disease resistance of Arabidopsis against Fusarium oxysporum after GbSBT1 overexpression.Wild-type (WT): Arabidopsis Columbia type Col-0; OEX: transgenic GbSBT1 lines. Left panels: un-inoculated plants; right panels: plants 2 days after F. oxysporum inoculation. (A–B) WT plants 0 and 2 days after ddH2O inoculation; (C–D) WT plants 0 and 2 days after F. oxysporum inoculation; (E–F) transgenic GbSBT1 plants 0 and 2 days after inoculation with ddH2O containing 0.2% Tween-20; (G–H) transgenic GbSBT1 plants 0 and 2 days after F. oxysporum inoculation. At least three biological replicates were performed in the F. oxysporum resistance analysis.
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pone.0153988.g006: Enhanced disease resistance of Arabidopsis against Fusarium oxysporum after GbSBT1 overexpression.Wild-type (WT): Arabidopsis Columbia type Col-0; OEX: transgenic GbSBT1 lines. Left panels: un-inoculated plants; right panels: plants 2 days after F. oxysporum inoculation. (A–B) WT plants 0 and 2 days after ddH2O inoculation; (C–D) WT plants 0 and 2 days after F. oxysporum inoculation; (E–F) transgenic GbSBT1 plants 0 and 2 days after inoculation with ddH2O containing 0.2% Tween-20; (G–H) transgenic GbSBT1 plants 0 and 2 days after F. oxysporum inoculation. At least three biological replicates were performed in the F. oxysporum resistance analysis.

Mentions: To better understand the molecular mechanisms of GbSBT1 in the immune response, we generated and self-crossed transgenic GbSBT1 Arabidopsis plants. After three rounds of self-crossing, two homologous lines showing different expression levels of the GbSBT1 gene were randomly selected and subjected to F. oxysporum and V. dahliae challenge. qPCR analysis of GbSBT1 expression levels in transgenic Arabidopsis lines OEX1 and OEX2 were carried out (S2 Fig). Obvious differences in disease symptoms between the WT and transgenic plants were observed at 2 days post-inoculation (dpi). The necrotic lesions were larger in the WT leaves than in the transgenic leaves (Fig 6). The disease spot diameter was also significantly smaller in the transgenic plants (0.36 ± 0.31 mm) than in the control plants (2.52 ± 0.66 mm). Similarly, necrotic lesions of V. dahliae on Arabidopsis leaves were smaller in the transgenic plants than in the WT plants (Fig 7). This result suggests that GbSBT1 overexpression inhibits the spread of necrotic lesions in transgenic leaves. Moreover, the control plants showed more pathogen aggregates near their leaf vessels, whereas the transgenic plants showed a negative result for trypan blue staining (Fig 8). As the infection progressed, the control plants wilted, whereas the transgenic GbSBT1 plants grew normally. These results demonstrate that the ectopic expression of GbSBT1 confers the increased tolerance of Arabidopsis against F. oxysporum infection.


Characterization of a Novel Cotton Subtilase Gene GbSBT1 in Response to Extracellular Stimulations and Its Role in Verticillium Resistance.

Duan X, Zhang Z, Wang J, Zuo K - PLoS ONE (2016)

Enhanced disease resistance of Arabidopsis against Fusarium oxysporum after GbSBT1 overexpression.Wild-type (WT): Arabidopsis Columbia type Col-0; OEX: transgenic GbSBT1 lines. Left panels: un-inoculated plants; right panels: plants 2 days after F. oxysporum inoculation. (A–B) WT plants 0 and 2 days after ddH2O inoculation; (C–D) WT plants 0 and 2 days after F. oxysporum inoculation; (E–F) transgenic GbSBT1 plants 0 and 2 days after inoculation with ddH2O containing 0.2% Tween-20; (G–H) transgenic GbSBT1 plants 0 and 2 days after F. oxysporum inoculation. At least three biological replicates were performed in the F. oxysporum resistance analysis.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4835097&req=5

pone.0153988.g006: Enhanced disease resistance of Arabidopsis against Fusarium oxysporum after GbSBT1 overexpression.Wild-type (WT): Arabidopsis Columbia type Col-0; OEX: transgenic GbSBT1 lines. Left panels: un-inoculated plants; right panels: plants 2 days after F. oxysporum inoculation. (A–B) WT plants 0 and 2 days after ddH2O inoculation; (C–D) WT plants 0 and 2 days after F. oxysporum inoculation; (E–F) transgenic GbSBT1 plants 0 and 2 days after inoculation with ddH2O containing 0.2% Tween-20; (G–H) transgenic GbSBT1 plants 0 and 2 days after F. oxysporum inoculation. At least three biological replicates were performed in the F. oxysporum resistance analysis.
Mentions: To better understand the molecular mechanisms of GbSBT1 in the immune response, we generated and self-crossed transgenic GbSBT1 Arabidopsis plants. After three rounds of self-crossing, two homologous lines showing different expression levels of the GbSBT1 gene were randomly selected and subjected to F. oxysporum and V. dahliae challenge. qPCR analysis of GbSBT1 expression levels in transgenic Arabidopsis lines OEX1 and OEX2 were carried out (S2 Fig). Obvious differences in disease symptoms between the WT and transgenic plants were observed at 2 days post-inoculation (dpi). The necrotic lesions were larger in the WT leaves than in the transgenic leaves (Fig 6). The disease spot diameter was also significantly smaller in the transgenic plants (0.36 ± 0.31 mm) than in the control plants (2.52 ± 0.66 mm). Similarly, necrotic lesions of V. dahliae on Arabidopsis leaves were smaller in the transgenic plants than in the WT plants (Fig 7). This result suggests that GbSBT1 overexpression inhibits the spread of necrotic lesions in transgenic leaves. Moreover, the control plants showed more pathogen aggregates near their leaf vessels, whereas the transgenic plants showed a negative result for trypan blue staining (Fig 8). As the infection progressed, the control plants wilted, whereas the transgenic GbSBT1 plants grew normally. These results demonstrate that the ectopic expression of GbSBT1 confers the increased tolerance of Arabidopsis against F. oxysporum infection.

Bottom Line: Verticillium pathogens secrete various disease-causing effectors in cotton.Moreover, the GbSBT1 protein is mainly localized in the cell membrane and moves into the cytoplasm following jasmonic acid and ethylene treatments.In summary, GbSBT1 recognizes the effector PHB protein secreted from V. dahliae and is involved in Verticillium-induced resistance in cotton.

View Article: PubMed Central - PubMed

Affiliation: Plant Biotechnology Research Center, School of Agriculture and Life Sciences, Shanghai Jiao Tong University, Shanghai, China.

ABSTRACT
Verticillium wilt is a disastrous vascular disease in plants caused by Verticillium dahliae. Verticillium pathogens secrete various disease-causing effectors in cotton. This study identified a subtilase gene GbSBT1 from Gossypium babardense and investigated the roles against V. dahliae infection. GbSBT1 gene expression is responsive to V. dahliae defense signals, jasmonic acid, and ethylene treatments. Moreover, the GbSBT1 protein is mainly localized in the cell membrane and moves into the cytoplasm following jasmonic acid and ethylene treatments. Silencing GbSBT1 gene expression through virus-induced GbSBT1 gene silencing reduced the tolerance of Pima-90 (resistant genotype), but not facilitated the infection process of V. dahliae in Coker-312 (sensitive genotype). Moreover, the ectopically expressed GbSBT1 gene enhanced the resistance of Arabidopsis to Fusarium oxysporum and V. dahliae infection and activated the expression levels of defense-related genes. Furthermore, pull-down, yeast two-hybrid assay, and BiFC analysis revealed that GbSBT1 interacts with a prohibitin (PHB)-like protein expressed in V. dahliae pathogens during infection. In summary, GbSBT1 recognizes the effector PHB protein secreted from V. dahliae and is involved in Verticillium-induced resistance in cotton.

No MeSH data available.


Related in: MedlinePlus