Abstract
Main conclusion
Root transcriptomics and biochemical analyses in water-stressed Pisum sativum plants inoculated with Pseudomonas spp. suggested preservation of ABA-related pathway and ROS detoxification, resulting in an improved tolerance to stress.
Abstract
Drought already affects agriculture in large areas of the globe and, due to climate change, these areas are predicted to become increasingly unsuitable for agriculture. For several years, plant growth-promoting bacteria (PGPB) have been used to improve legume yields, but many aspects of this interaction are still unclear. To elucidate the mechanisms through which root-associated PGPB can promote plant growth in dry environments, we investigated the response of pea plants inoculated with a potentially beneficial Pseudomonas strain (PK6) and subjected to two different water regimes. Combined biometric, biochemical, and root RNA-seq analyses revealed that PK6 improved pea growth specifically under water deficit, as inoculated plants showed an increased biomass, larger leaves, and longer roots. Abscisic acid (ABA) and proline quantification, together with the transcriptome analysis, suggested that PK6-inoculated plant response to water deficit was more diversified compared to non-inoculated plants, involving alternative metabolic pathways for the detoxification of reactive oxygen species (ROS) and the preservation of the ABA stress signaling pathway. We suggest that the metabolic response of PK6-inoculated plants was more effective in their adaptation to water deprivation, leading to their improved biometric traits. Besides confirming the positive role that PGPB can have in the growth of a legume crop under adverse conditions, this study offers novel information on the mechanisms regulating plant–bacteria interaction under varying water availability. These mechanisms and the involved genes could be exploited in the future for the development of legume varieties, which can profitably grow in dry climates.
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Data availability
The data supporting the findings of this study are available in the Supplementary Information of this article. Reads from RNA-seq were submitted to NCBI Sequence Read Archive (SRA) under Bioproject accession number PRJNA980113.
Abbreviations
- DAT:
-
Days after the transplant
- DEG:
-
Differentially expressed gene
- FC:
-
Fold change
- GO:
-
Gene ontology
- NI:
-
Non-inoculated
- PGPB:
-
Plant growth-promoting bacteria
- POD:
-
Peroxidase
- ROS:
-
Reactive oxygen species
- TRL:
-
Total root length
- WR:
-
Water regime
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Acknowledgements
The authors are grateful to Dr. Raffaella Balestrini for the useful comments and discussion on the study, as well as for critically reviewing the manuscript.
Funding
The work was supported by the National Research Council of Italy — PMAS Arid Agriculture University Rawalpindi-Pakistan Joint Laboratory Project 2021–2023 “Selection and evaluation of plant growth promoting rhizobacteria to increase climate-resilient crop production” (PI: Dr. Mauro Centritto).
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Conceptualization: FS; methodology: MS, FS, EZ, CB, and AG; formal analysis and investigation: MS, FS, and EZ; writing—original draft preparation: MS and EZ; writing—review and editing: MS, FS, and EZ;
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Supplementary file 1. Fig. S1
: Shoot length of pea plants recorded from 12 to 33 days after the transplant (DAT). NI, non-inoculated; PK6, PK6-inoculated; WR1, water regime 1; WR2, water regime 2. Symbols represent the mean shoot length of each treatment; n = 5 for NIWR1, NIWR2, n = 4 for PK6WR1 and PK6WR2. (JPG 837 kb)
Supplementary file 2. Fig. S2
: Leaf number and % of wilted leaves. NI, non-inoculated; PK6, PK6-inoculated; WR1, water regime 1; WR2, water regime 2; n = 5 for NIWR1 and NIWR2, n = 4 for PK6WR1 and PK6WR2. Individual standard deviations are used to calculate the intervals. Bars represent the standard error of the mean. Above the intervals, different letters display statistical differences among the treatments (Fisher LSD pairwise comparison, 95% confidence). (JPG 120 kb)
Supplementary file 3. Fig. S3
: Concentration (%) of C and N in dry shoots of pea. NI, non-inoculated; WR1, water regime 1; WR2, water regime 2; DW, dry weight; n = 5 for NIWR1 and NIWR2, n = 4 for PK6WR1 and PK6WR2. Individual standard deviations are used to calculate the intervals. Bars represent the standard error of the mean. Above the intervals, different letters display statistical differences among the treatments (Fisher LSD pairwise comparison, 95% confidence). (JPG 111 kb)
Supplementary file 4. Fig. S4
: Correlation plot between expression data obtained by RNA-seq (log2FC by RNA-seq) and RT-qPCR analysis (log2FC by RT-qPCR). Regression line (dark grey line), R-square and associated P-value are shown. The gray shadow represents the 95% confidence interval. Tested genes are labelled as in Table S5. NI, non-inoculated; PK6, PK6-inoculated; WR1, water regime 1; WR2, water regime 2. (JPG 1297 kb)
Supplementary file 5. Fig. S5
: Heatmap displaying the relative expression values (log2FC, comparison between WR2 and WR1) of genes involved in the GO names “proline metabolic process” and “hydrogen peroxide catabolic process”. Genes in white were not significantly up- or down-regulated (Wald test, P-value corrected for multiple testing > 0.05). NI, non-inoculated; PK6, PK6-inoculated; WR1, water regime 1; WR2, water regime 2. (JPG 755 kb)
Supplementary file 6. Fig. S6
: Heatmap displaying the relative expression values (log2FC, comparison between WR2 and WR1) of genes involved in the GO names “response to heat”. Genes in white were not significantly up- or down-regulated (Wald test, P-value corrected for multiple testing > 0.05). NI, non-inoculated; PK6, PK6-inoculated; WR1, water regime 1; WR2, water regime 2. (JPG 579 kb)
Supplementary file 7. Fig. S7
: Heatmap displaying the relative expression values (log2FC, comparison between WR2 and WR1) of genes involved in the GO names “coumarin metabolic process” and “phenylpropanoid metabolic process”. Genes in white were not significantly up- or down-regulated (Wald test, P-value corrected for multiple testing > 0.05). NI, non-inoculated; PK6, PK6-inoculated; WR1, water regime 1; WR2, water regime 2. (JPG 720 kb)
Supplementary file 8. Fig. S8
: Heatmap displaying the relative expression values (log2FC, comparison between WR2 and WR1) of genes involved in the GO names “biological process involved in interspecies interaction between organisms” and “response to fungus”. Genes in white were not significantly up- or down-regulated (Wald test, P-value corrected for multiple testing > 0.05). NI, non-inoculated; PK6, PK6-inoculated; WR1, water regime 1; WR2, water regime 2. (JPG 1333 kb)
Supplementary file 9. Fig. S9
: Heatmap displaying the relative expression values (log2FC, comparison between WR2 and WR1) of genes involved in the GO names “regulation of root hair elongation” and “regulation of leaf senescence”. Genes in white were not significantly up- or down-regulated (Wald test, P-value corrected for multiple testing > 0.05). NI, non-inoculated; PK6, PK6-inoculated; WR1, water regime 1; WR2, water regime 2. (JPG 122 kb)
Supplementary file 10. Fig. S10
: Heatmap displaying the relative expression values (log2FC, comparison between WR2 and WR1) of genes putatively involved in ABA metabolism. Genes in white were not significantly up- or down-regulated (Wald test, P-value corrected for multiple testing > 0.05). (JPG 2022 kb)
Supplementary file 11. Table S1
: Characteristics of primers used in the RT-qPCR analysis on selected DEGs. Transcript sequence IDs are from P. sativum Ensembl database. (XLSX 7 kb)
Supplementary file 12. Table S2
: Summary Statistics of RNA-Seq alignment of pea roots reads. NI, non-inoculated; PK6, PK6-inoculated; WR1, water regime 1; WR2, water regime 2. (XLSX 6 kb)
Supplementary file 13. Table S3
: Raw RNA-seq pea root reads count (TPM). NI, non-inoculated; PK6, PK6-inoculated; WR1, water regime 1; WR2, water regime 2. (XLSX 3892 kb)
Supplementary file 14. Table S4
: Significant (P adj < 0.05) DEGs obtained comparing the NIWR1, NIWR2, PK6WR1 and PK6WR2 treatments. NI, non-inoculated; PK6, PK6-inoculated; WR1, water regime 1; WR2, water regime 2. (XLSX 691 kb)
Supplementary file 15. Table S5
: RT-qPCR gene expression data of five up-regulated DEGs, five down-regulated DEGs and three DEGs putatively coding for peroxidases obtained in the comparison between WR2 and WR1 plants. NI, non-inoculated; PK6, PK6-inoculated; WR1, water regime 1; WR2, water regime 2. (XLSX 6 kb)
Supplementary file 16. Table S6
: Fold changes of DEGs assigned to GO enriched terms. DEGs in blue font are from the comparison NIWR2 versus NIWR1, DEGs in red font are from the comparison PK6WR2 versus PK6WR1. NI, non-inoculated; PK6, PK6-inoculated; WR1, water regime 1; WR2, water regime 2. (XLSX 126 kb)
Supplementary file 17. Table S7
: Fold changes of genes related to ABA metabolism in the comparisons NIWR2 versus NIWR1 and PK6WR2 versus PK6WR1. NI, non-inoculated; PK6, PK6-inoculated; WR1, water regime 1; WR2, water regime 2. (XLSX 12 kb)
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Schillaci, M., Zampieri, E., Brunetti, C. et al. Root transcriptomic provides insights on molecular mechanisms involved in the tolerance to water deficit in Pisum sativum inoculated with Pseudomonas sp.. Planta 259, 33 (2024). https://doi.org/10.1007/s00425-023-04310-0
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DOI: https://doi.org/10.1007/s00425-023-04310-0