Insights into the mechanisms implicated in Pinus pinaster resistance to pinewood nematode
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Date
Authors
Modesto, Ines
Sterck, Lieven
Arbona, Vicent
Gomez-Cadenas, Aurelio
Carrasquinho, Isabel
Van de Peer, Yves
Miguel, Celia M.
Journal Title
Journal ISSN
Volume Title
Publisher
Frontiers Media
Abstract
Pine wilt disease (PWD), caused by the plant–parasitic nematode Bursaphelenchus
xylophilus, has become a severe environmental problem in the Iberian Peninsula
with devastating effects in Pinus pinaster forests. Despite the high levels of this
species’ susceptibility, previous studies reported heritable resistance in P. pinaster trees.
Understanding the basis of this resistance can be of extreme relevance for future
programs aiming at reducing the disease impact on P. pinaster forests. In this study,
we highlighted the mechanisms possibly involved in P. pinaster resistance to PWD, by
comparing the transcriptional changes between resistant and susceptible plants after
infection. Our analysis revealed a higher number of differentially expressed genes (DEGs)
in resistant plants (1,916) when compared with susceptible plants (1,226). Resistance
to PWN is mediated by the induction of the jasmonic acid (JA) defense pathway,
secondary metabolism pathways, lignin synthesis, oxidative stress response genes,
and resistance genes. Quantification of the acetyl bromide-soluble lignin confirmed a
significant increase of cell wall lignification of stem tissues around the inoculation zone
in resistant plants. In addition to less lignified cell walls, susceptibility to the pine wood
nematode seems associated with the activation of the salicylic acid (SA) defense pathway
at 72 hpi, as revealed by the higher SA levels in the tissues of susceptible plants. Cell
wall reinforcement and hormone signaling mechanisms seem therefore essential for a
resistance response.
Description
Supplementary Figure 1. Boxplots of the height and diameter at the base of the
stem of inoculated plants (half-sib family 440) and t-test results for the comparison
of these parameters’ means between resistant (res) and susceptible (sus) plants.
(A) Boxplot of height (cm) measurements. (B) Boxplot of diameter at the base of
the stem (mm) measurements. Both measurements were made before inoculations. (C) t-Test results for heights comparison. (D) t-Test results for
diameter comparison. N, number of samples. SD, standard deviation.
Supplementary Figure 2. Heatmaps representing the expression patterns of genes involved in secondary metabolism. (A) Flavonoid biosynthesis pathway. (B) Terpenoid biosynthesis pathways, including terpenoid backbone biosynthesis (Terp. Backbone), monoterpenoid biosynthesis, sesquiterpenoid biosynthesis, and diterpenoid biosynthesis pathways. The color gradient represents mean expression levels (logTPM) of each gene for control (C), susceptible (S), and resistant (R) samples.
Supplementary Figure 3. Heatmaps representing the expression patterns of genes involved in the synthesis of hydrogen peroxide (A) and response to oxidative stress (B). The color gradient represents mean expression levels (logTPM) of each gene for control (C), susceptible (S), and resistant (R) samples.
Supplementary Figure 4. Heatmaps representing the expression patterns of hormone responsive transcription factors (TFs). Jasmonate responsive TFs JAZ/Tify (A) and ERF (B), salicylic acid responsive TFs WRKY (C), and abscisic acid responsive TFs NAC (D). The color gradient represents mean expression levels (logTPM) of each gene for control (C), susceptible (S), and resistant (R) samples.
Supplementary Table 1. De novo assembly and P. pinaster reference transcriptome statistics.
Supplementary Table 2. Genes selected for quantitative RT-qPCR, respective primer sequences, amplicon size, and annealing temperatures used.
Supplementary Table 3. Genes expressed by Bursaphelenchus xylophilus in inoculated samples.
Supplementary Table 4. Differential expressed genes in susceptible plants when compared to controls (log2 fold change ≥ |2|, FDR corrected p-value (padj) ≤ 0.05). InterPro, KEGG, and blastx annotations are presented.
Supplementary Table 5. Differential expressed genes in resistant plants when compared to controls (log2 fold change ≥ |2|, FDR corrected p-value (padj) ≤ 0.05). InterPro, KEGG, and blastx annotations are presented.
Supplementary Table 6. GO terms enriched in the upregulated genes in susceptible samples when compared with controls, after trimming for redundancy.
Supplementary Table 7. GO terms enriched in the upregulated genes in resistant samples when compared with controls, after trimming for redundancy.
Supplementary Table 8. Complete list of genes annotated with DRAGO 2 tool.
Supplementary Table 9. Genes used for calculating average log2(fold change) expression levels for Figures 5, 6.
Data Sheet 1. Fasta file with de novo assembled Pinus pinaster transcripts.
Supplementary Figure 2. Heatmaps representing the expression patterns of genes involved in secondary metabolism. (A) Flavonoid biosynthesis pathway. (B) Terpenoid biosynthesis pathways, including terpenoid backbone biosynthesis (Terp. Backbone), monoterpenoid biosynthesis, sesquiterpenoid biosynthesis, and diterpenoid biosynthesis pathways. The color gradient represents mean expression levels (logTPM) of each gene for control (C), susceptible (S), and resistant (R) samples.
Supplementary Figure 3. Heatmaps representing the expression patterns of genes involved in the synthesis of hydrogen peroxide (A) and response to oxidative stress (B). The color gradient represents mean expression levels (logTPM) of each gene for control (C), susceptible (S), and resistant (R) samples.
Supplementary Figure 4. Heatmaps representing the expression patterns of hormone responsive transcription factors (TFs). Jasmonate responsive TFs JAZ/Tify (A) and ERF (B), salicylic acid responsive TFs WRKY (C), and abscisic acid responsive TFs NAC (D). The color gradient represents mean expression levels (logTPM) of each gene for control (C), susceptible (S), and resistant (R) samples.
Supplementary Table 1. De novo assembly and P. pinaster reference transcriptome statistics.
Supplementary Table 2. Genes selected for quantitative RT-qPCR, respective primer sequences, amplicon size, and annealing temperatures used.
Supplementary Table 3. Genes expressed by Bursaphelenchus xylophilus in inoculated samples.
Supplementary Table 4. Differential expressed genes in susceptible plants when compared to controls (log2 fold change ≥ |2|, FDR corrected p-value (padj) ≤ 0.05). InterPro, KEGG, and blastx annotations are presented.
Supplementary Table 5. Differential expressed genes in resistant plants when compared to controls (log2 fold change ≥ |2|, FDR corrected p-value (padj) ≤ 0.05). InterPro, KEGG, and blastx annotations are presented.
Supplementary Table 6. GO terms enriched in the upregulated genes in susceptible samples when compared with controls, after trimming for redundancy.
Supplementary Table 7. GO terms enriched in the upregulated genes in resistant samples when compared with controls, after trimming for redundancy.
Supplementary Table 8. Complete list of genes annotated with DRAGO 2 tool.
Supplementary Table 9. Genes used for calculating average log2(fold change) expression levels for Figures 5, 6.
Data Sheet 1. Fasta file with de novo assembled Pinus pinaster transcripts.
Keywords
Cell wall lignification, Jasmonate, Maritime pine, Resistance genes, Secondary metabolism, Transcriptome, Bursaphelenchus xylophilus, Pine wilt disease (PWD), Pinus pinaster, Differentially expressed genes (DEGs)
Sustainable Development Goals
Citation
Modesto, I., Sterck, L., Arbona, V., Gomez-Cadenas, A., Carrasquinho, I., Van de Peer, Y. &. Miguel, C.M. (2021)
Insights Into the Mechanisms
Implicated in Pinus pinaster
Resistance to Pinewood Nematode.
Frontiers in Plant Science 12:690857.
DOI: 10.3389/fpls.2021.690857.