Resistance to hemi-biotrophic F. graminearum infection is associated with coordinated and ordered expression of diverse defense signaling pathways

Abstract

Fusarium species cause serious diseases in cereal staple food crops such as wheat and maize. Currently, the mechanisms underlying resistance to Fusarium-caused diseases are still largely unknown. In the present study, we employed a combined proteomic and transcriptomic approach to investigate wheat genes responding to F. graminearum infection that causes Fusarium head blight (FHB). We found a total of 163 genes and 37 proteins that were induced by infection. These genes and proteins were associated with signaling pathways mediated by salicylic acid (SA), jasmonic acid (JA), ethylene (ET), calcium ions, phosphatidic acid (PA), as well as with reactive oxygen species (ROS) production and scavenging, antimicrobial compound synthesis, detoxification, and cell wall fortification. We compared the time-course expression profiles between FHB-resistant Wangshuibai plants and susceptible Meh0106 mutant plants of a selected set of genes that are critical to the plants' resistance and defense reactions. A biphasic phenomenon was observed during the first 24 h after inoculation (hai) in the resistant plants. The SA and Ca(2+) signaling pathways were activated within 6 hai followed by the JA mediated defense signaling activated around 12 hai. ET signaling was activated between these two phases. Genes for PA and ROS synthesis were induced during the SA and JA phases, respectively. The delayed activation of the SA defense pathway in the mutant was associated with its susceptibility. After F. graminearum infection, the endogenous contents of SA and JA in Wangshuibai and the mutant changed in a manner similar to the investigated genes corresponding to the individual pathways. A few genes for resistance-related cell modification and phytoalexin production were also identified. This study provided important clues for designing strategies to curb diseases caused by Fusarium.

Figure 1. Disease symptoms of Wangshuibai and the susceptible Meh0106 mutant 15 d after F. graminearum inoculation, compared with water-mimicked treatment.

Scale bar represents 2.5 cm.

Figure 2. Silver-stained 2-DE of proteins extracted from Wangshuibai and Meh0106 spikes at 12 hai with F. graminearum or H₂O.

This is a representative image from three technical and three biological replicates.

Figure 3. Venn diagram indicating number of proteins differentially expressed in Wangshuibai (WSB) and Meh0106 after F. graminearum infection.

A. Number of up-regulated proteins; B. Number of down-regulated proteins.

Figure 4. Percentage of the Arabidopsis homologs up-regulated by biotic stresses.

Figure 5. Expression profiles of PAL (A), Glu2 (B), ICS1 (C), Ca²⁺ ATPase (D), CaM (E), DGK (F), PLD (G), ACO (H), 12-OPR3 (I), ERF1 (J), PR3 (K), EDS1 (L), MYC2 (M), BIK1 (N), NPR1 (O), WRKY33 (P), MKP-1 (Q), NADPH oxidase (R), SAMDC (S), PAO (T), CCOMT (U), CHS (V), CYP71D8 (W) and Chitinase 1 (X) in Wangshuibai (WSB) and Meh0106 spikes after F. graminearum infection.

The expression levels were relative to no inoculation (0 h) after normalization of the qRT-PCR outputs with the wheat tubulin gene output. RT-PCR was performed using gene-specific primers (Table S4). The experiment was repeated three times with similar results. Data were presented as average + S.D with n = 3.

Figure 6. Activities of SOD (A) and POD (B) after F. graminearum infection in Wangshuibai and Meh0106 spikes.

Each data point consisted of three replicates. Error bars indicate SD. WSB: Wangshuibai; Mutant: Meh0106.

Figure 7. Endogenous contents of SA (A) and JA (B) in Wangshuibai and Meh0106 spikes at different time points after F. graminearum infection.

Each data point consisted of three replicates. The experiments were repeated twice with similar results. Error bars indicate SD.

Figure 8. A model illustrating pathways leading to FHB resistance.

PIs, protease inhibitors; XET, xyloglucan endotransglycosylase; Xls, xylanase inhibitors. See the text for other abbreviations. The peripheral solid lines indicate cell wall, the peripheral dotted lines indicate plasma membrane; the solid arrows represent direct interactions, and the dotted arrows represent indirect interactions. Perpendicular line on arrows indicates negative relationship.