Multiple phytohormone signalling pathways modulate susceptibility of tomato plants to Alternaria alternata f. sp. lycopersici

Abstract

Three phytohormone molecules - ethylene (ET), jasmonic acid (JA) and salicylic acid (SA) - play key roles in mediating disease response to necrotrophic fungal pathogens. This study investigated the roles of the ET, JA, and SA pathways as well as their crosstalk during the interaction between tomato (Solanum lycopersicum) plants and a necrotrophic fungal pathogen Alternaria alternata f. sp. lycopersici (AAL). Both the ET and JASMONIC ACID INSENSITIVE1 (JAI1) receptor-dependent JA signalling pathways are necessary for susceptibility, while SA response promotes resistance to AAL infection. In addition, the role of JA in susceptibility to AAL is partly dependent on ET biosynthesis and perception, while the SA pathway enhances resistance to AAL and antagonizes the ET response. Based on these results, it is proposed that ET, JA, and SA each on their own can influence the susceptibility of tomato to AAL. Furthermore, the functions of JA and SA in susceptibility to the pathogen are correlated with the enhanced or decreased action of ET, respectively. This study has revealed the functional relationship among the three key hormone pathways in tomato defence against AAL.

Fig. 3.

The role of the SA pathway in tomato resistance to A. alternata f. sp. lycopersici. (A) Effects of various concentrations of SA application on the disease development in CM plants. (B) Disease development in the NahG transgenic line and MM plants. (C) Fungal biomass in MM and NahG transgenic plants at 6 d post inoculation. Error bars indicate standard deviation from the mean of three replicates. In A, asterisks indicate significant differences as compared with the water-treated control at the same time point (P < 0.05; Student’s t-test), the experiment was repeated twice and similar results were obtained; for each time point in B, letters indicate significant differences among treatments (P < 0.05, Duncan’s multiple range test); in C, letters indicate significant differences among treatments (P < 0.05, Duncan’s multiple range test). (D, E) Transcription patterns of SA-regulated PR genes PR1a and PR2a, respectively, in MM and NahG plants at 0, 1, 3, and 5 d post inoculation.

Fig. 1.

Role of ET in tomato susceptibility to A. alternata f. sp. lycopersici. (A) Effect of exogenous ET precursor ACC (0.1mM) and ET action inhibitor silver thiosulphate (1mM) on disease development in CM plants. (B) Effect of ET receptor inhibitor MCP (10 nl l^–1) on the disease development in CM plants. (C) Effect of 10 nl l^–1 1-MCP and 0.1mM ACC on the relative fungal DNA in CM leaves at 6 d post inoculation; the amount of fungal DNA was quantified by qPCR using ALT1 primers. (D) Disease symptoms in epi and VFN8 plants at 7 d post inoculation. Error bars indicate standard deviation from the mean of three replicates. For each time point in A and B, letters indicate significant differences among treatments; in C, letters indicate significant differences among treatments (P < 0.05, Duncan’s multiple range test). The experiment was repeated three times and similar results were obtained. For each experiment, data were analysed separately. Results of one representative experiment are shown.

Fig. 2.

The role of the JA pathway in tomato susceptibility to A. alternata f. sp. lycopersici. (A, B) Disease development in CM, spr2, jai1, and 35S::prosys plants (A) and effect of application of MeJA to CM and spr2 plants (B) at 4, 6, and 8 d post inoculation. (C) Fungal biomass in CM, spr2, jai1, and 35S::prosys plants at 6 d post inoculation. The amount of fungal DNA was quantified by qPCR using ALT1 gene primers. Error bars indicate standard deviation from the mean of three replicates. For each time point in A and B, letters indicate significant differences among treatments; in C, letters indicate significant differences among treatments (P < 0.05, Duncan’s multiple range test).

Fig. 4.

Transcription patterns of ET-related genes and ET production in jai1 and CM plants in response to A. alternata f. sp. lycopersici. Transcription patterns of ACS4 (A) ACO1 (B), NR (D), ETR4 (E), and ERF1 (F) in CM and jai1 plants at 0, 1, 3, and 5 d post infection. (C) ET production in jai1 and CM leaves during infection in a time-course experiment.

Fig. 5.

Effects of ET response to JA mutants or 35S::prosys and MeJA application to ET-biosynthetic or -insensitive tomato plants on susceptibility to A. alternata f. sp. lycopersici. (A) The effect of exogenous ET precursor ACC on spr2 and ET action inhibitor silver thiosulphate on 35S::prosys susceptibility to infection. (B) The effect of ACC on jai1 susceptibility to infection. (C) Fungal biomass in CM, jai1, and ACC-pretreated jai1 plants at 6 d post inoculation. (D) The effect of exogenous MeJA application to ET-biosynthetic or -insensitive tomato plants on disease development. Error bars indicate standard deviation from the mean of three replicates. For each time point in A, B, and D, letters indicate significant differences among treatments; in C, letters indicate significant differences among treatments (P < 0.05, Duncan’s multiple range test). The experiment was repeated once and similar results were obtained.

Fig. 6.

Effects of ET action inhibitor silver thiosulphate application to NahG transgenic line (A) and SA treatment of epi mutant (B) on disease development in tomato plants. Error bars indicate standard deviation from the mean of three replicates. In A, letters indicate significant differences among treatments (P < 0.05, Duncan’s multiple-range test). The experiment was repeated once and similar results were obtained. For each experiment, data were analysed separately. Results of one representative experiment are shown.

Fig. 7.

Transcription patterns of ET-biosynthetic genes ACS2 and ACO1 and ET-regulated genes NR, ETR4, and ERF1 in NahG and MM plants in response to A. alternata f. sp. lycopersici. Transcript accumulation of ACS2 (A), ACO1 (B), NR (C), ETR4 (D), and ERF1 (E) in A. alternata f. sp. lycopersici-infected MM and NahG plants at 0, 1, 3, and 5 d post infection.

Fig. 8.

A proposed model for the JA, SA, and ET defence signalling pathways and their interactions during A. alternata f. sp. lycopersici infection. Both ET and JAI1 receptor-dependent JA pathways are necessary for susceptibility, while SA signalling promotes resistance of tomato plants to infection. Furthermore, the JA and ET pathways act synergistically to promote the susceptibility, while SA promotes tomato resistance to AAL and antagonizes ET signalling during AAL infection. JA and SA affect tomato susceptibility to the pathogen partly through the action of ET. ET-mediated signalling plays a central role in determining the susceptibility of tomato to A. alternata f. sp. lycopersici.