Phospholipid production and signaling by a plant defense inducer against Podosphaera xanthii is genotype-dependent

Abstract

Biotrophic phytopathogenic fungi such as Podosphaera xanthii have evolved sophisticated mechanisms to adapt to various environments causing powdery mildews leading to substantial yield losses. Today, due to known adverse effects of pesticides, development of alternative control means is crucial and can be achieved by combining plant protection products with resistant genotypes. Using plant defense inducers, natural molecules that stimulate plant immune system mimicking pathogen attack is sustainable, but information about their mode of action in different hosts or host genotypes is extremely limited. Reynoutria sachalinensis extract, a known plant defense inducer, especially through the Salicylic acid pathway in Cucurbitaceae crops against P. xanthii, was employed to analyze the signaling cascade of defense activation. Here, we demonstrate that R. sachalinensis extract enhances phospholipid production and signaling in a Susceptible to P. xanthii courgette genotype, while limited response is observed in an Intermediate Resistance genotype due to genetic resistance. Functional enrichment and cluster analysis of the upregulated expressed genes revealed that inducer application promoted mainly lipid- and membrane-related pathways in the Susceptible genotype. On the contrary, the Intermediate Resistance genotype exhibited elevated broad spectrum defense pathways at control conditions, while inducer application did not promote any significant changes. This outcome was obvious and at the metabolite level. Main factor distinguishing the Intermediate Resistance form the Susceptible genotype was the epigenetic regulated increased expression of a G3P acyltransferase catalyzing phospholipid production. Our study provides evidence on phospholipid-based signaling after plant defense inducer treatment, and the selective role of plant’s genetic background.

Author Contributions

TM and EM designed the research. TM performed most of the experiments. EB, CA and KEV contributed in data collection and analysis tools. TM wrote the original draft. All authors reviewed and approved the final version of the manuscript.

Data availability

The data underlying this article are available in the article and its online supplementary material.

Conflict of interest statement

The authors declare that there is no conflict of interest.

Supplementary Data

Supplementary data is available at Horticulture Research online.

References

[1] Burketova L, Trda L, Ott PG et al. Bio-based resistance inducers for sustainable plant protection against pathogens. Biotechnol Adv. 2015; 33:994-1004.

[2] W.H.O. Public Health Impact of Pesticides Used in Agriculture. Geneva, Switzerland: World Health Organization, 1990.

[3] Zubrod JP, Bundschuh M, Arts G et al. Fungicides: an overlooked pesticide class? Environ Sci Technol. 2019; 53:3347-65.

[4] Fisher MC, Henk DA, Briggs CJ et al. Emerging fungal threats to animal, plant and ecosystem health. Nature. 2012; 484:186-94.

[5] EPPO. PP 1/319 (1) general principles for efficacy evaluation of plant protection products with a mode of action as plant defence inducers. Bulletin OEPP/EPPO Bulletin. 2021; 51:5-9.

[6] Jones JD, Dangl JL. The plant immune system. Nature. 2006; 444:323-9.

[7] Bari R, Jones JD. Role of plant hormones in plant defence responses. Plant Mol Biol. 2009; 69:473-88.

[8] Pieterse CM, Van der Does D, Zamioudis C et al. Hormonal modulation of plant immunity. Annu Rev Cell Dev Biol. 2012; 28:489-521.

[9] Macabuhay A, Arsova B, Walker R et al. Modulators or facilitators? Roles of lipids in plant root-microbe interactions. Trends Plant Sci. 2022; 27:180-90.

[10] Malinovsky FG, Fangel JU, Willats WG. The role of the cell wall in plant immunity. Front Plant Sci. 2014; 5:178.

[11] Boller T, Felix G. A renaissance of elicitors: perception of microbe-associated molecular patterns and danger signals by pattern-recognition receptors. Annu Rev Plant Biol. 2009; 60:379-406.

[12] Elmore JM, Coaker G. The role of the plasma membrane H⁺-ATPase in plant-microbe interactions. Mol Plant. 2011; 4:416-27.

[13] Tayeh C et al. Lipids as Markers of Induced Resistance in Wheat: A Biochemical and Molecular Approach in Lipid Metabolism 363-390. London, United Kingdom: IntechOpen, 2013.

[14] Cassim AM et al. Plant lipids: key players of plasma membrane organization and function. Prog Lipid Res. 2019; 73:1-27.

[15] Shah J. Lipids, lipases, and lipid-modifying enzymes in plant disease resistance. Annu Rev Phytopathol. 2005; 43:229-60.

[16] Hong Y, Zhao J, Guo L et al. Plant phospholipases D and C and their diverse functions in stress responses. Prog Lipid Res. 2016; 62:55-74.

[17] Xing J, Zhang L, Duan Z et al. Coordination of phospholipid-based signaling and membrane trafficking in plant immunity. Trends Plant Sci. 2021; 26:407-20.

[18] Canonne J, Froidure-Nicolas S, Rivas S. Phospholipases in action during plant defense signaling. Plant Signal Behav. 2011; 6:13-8.

[19] Eurostat G. & Components, M. Available online: https://ec.europa.eu/eurostat/databrowser/view.NAMQ_10_GDP/default/table (2020).

[20] Margaritopoulou T, Kizis D, Kotopoulis D et al. Enriched H3K4me3 marks at Pm-0 resistance-related genes prime courgette against Podosphaera xanthii. Plant Physiol. 2022; 188:576-92.

[21] Margaritopoulou T, Toufexi E, Kizis D et al. Reynoutria sachalinensis extract elicits SA-dependent defense responses in courgette genotypes against powdery mildew caused by Podosphaera xanthii. Sci Rep. 2020; 10:1-17.

[22] Shannon P, Markiel A, Ozier O et al. Cytoscape: a software environment for integrated models of biomolecular interaction networks. Genome Res. 2003; 13:2498-504.

[23] Szklarczyk D, Morris JH, Cook H et al. The STRING database in 2017:quality-controlled protein-protein association networks, made broadly accessible. Nucleic Acids Res. 2017; 45:D362-8.

[24] Li-Beisson Y, Shorrosh B, Beisson F et al. Acyl-lipid metabolism. Arabidopsis Book. 2013; 11:1-70.

[25] Stillwell W. Introduction to Biological Membranes.In: An Introduction to Biological Membranes. Elsevier, 2016, 3-15.

[26] Holdsworth WL, LaPlant KE, Bell DC et al. Cultivar-based introgression mapping reveals wild species-derived Pm-0, the major powdery mildew resistance locus in squash. PLoS One. 2016; 11:e0167715.

[27] Nishizawa-Yokoi A, Yoshida E, Yabuta Y et al. Analysis of the regulation of target genes by an Arabidopsis heat shock transcription factor, HsfA2. Biosci Biotechnol Biochem. 2009; 73:890-5.

[28] Chen G, Snyder CL, Greer MS et al. Biology and biochemistry of plant phospholipases. Crit Rev Plant Sci. 2011; 30:239-58.

[29] Mansfeld J, Gebauer S, et al. Dathe K Secretory phospholipase A2 from Arabidopsis thaliana: insights into the three-dimensional structure and the amino acids involved in catalysis. Biochemistry. 2006; 45:5687-94.

[30] Ryu SB. Phospholipid-derived signaling mediated by phospholipase a in plants. Trends Plant Sci. 2004; 9:229-35.

[31] Lee B-H, Choi SH, Kim HJ et al. Plant lysophosphatidic acids: a rich source for bioactive lysophosphatidic acids and their pharmacological applications. Biol Pharm Bull. 2016; 39:156-62.

[32] Zhao J. Phospholipase D and phosphatidic acid in plant defence response: from protein-protein and lipid-protein interactions to hormone signalling. J Exp Bot. 2015; 66:1721-36.

[33] Ali U, Lu S, Fadlalla T et al. The functions of phospholipases and their hydrolysis products in plant growth, development and stress responses. Prog Lipid Res. 2022; 86:101158.

[34] Saile SC, Ackermann FM, Sunil S et al. Arabidopsis ADR1 helper NLR immune receptors localize and function at the plasma membrane in a phospholipid dependent manner. New Phytol. 2021; 232:2440-56.

[35] Laxalt AM, Munnik T. Phospholipid signalling in plant defence. Curr Opin Plant Biol. 2002; 5:332-8.

[36] Donaldson JG. Phospholipase D in endocytosis and endosomal recycling pathways. Biochimica Et Biophysica Acta (BBA)-Molecular and Cell Biology of Lipids. 2009; 1791:845-9.

[37] Ohlrogge J, Browse J. Lipid biosynthesis. Plant Cell. 1995; 7:957-70.

[38] Kunst L, Browse J, Somerville C. Altered regulation of lipid biosynthesis in a mutant of Arabidopsis deficient in chloroplast glycerol-3-phosphate acyltransferase activity. Proc Natl Acad Sci. 1988; 85:4143-7.

[39] Xu C, Yu B, Cornish AJ et al. Phosphatidylglycerol biosynthesis in chloroplasts of Arabidopsis mutants deficient in acyl-ACP glycerol-3-phosphate acyltransferase. Plant J. 2006; 47:296-309.

[40] Chen X, Snyder CL, Truksa M et al. sn-Glycerol-3-phosphate acyltransferases in plants. Plant Signal Behav. 2011; 6:1695-9.

[41] Oerke E-C, Dehne H-W. Safeguarding production—losses in major crops and the role of crop protection. Crop Prot. 2004; 23:275-85.

[42] Pérez-García A, Romero D, Fernández-Ortuño D et al. The powdery mildew fungus Podosphaera fusca (synonym Podosphaera xanthii), a constant threat to cucurbits. Mol Plant Pathol. 2009; 10:153-60.

[43] Micali C, Göllner K, Humphry M et al. The powdery mildew disease of Arabidopsis: a paradigm for the interaction between plants and biotrophic fungi. Arabidopsis Book. 2008; 6:e0115.

[44] Daayf F, Schmitt A, Belanger R. The effects of plant extracts of Reynoutria sachalinensis on powdery mildew development and leaf physiology of long English cucumber. Plant Dis. 1995; 79:577-80.

[45] Daayf F, Schmitt A, Bélanger RR. Evidence of phytoalexins in cucumber leaves infected with powdery mildew following treatment with leaf extracts of Reynoutria sachalinensis. Plant Physiol. 1997; 113:719-27.

[46] Daayf F, Ongena M, Boulanger R et al. Induction of phenolic compounds in two cultivars of cucumber by treatment of healthy and powdery mildew-infected plants with extracts of Reynoutria sachalinensis. J Chem Ecol. 2000; 26:1579-93.

[47] Seth T, Asija S, Umar S et al. The intricate role of lipids in orchestrating plant defense responses. Plant Sci. 2024; 338:111904.

[48] Mehta S, Chakraborty A, Roy A et al. Fight hard or die trying: current status of lipid signaling during plant-pathogen interaction. Plan Theory. 2021; 10:1098.

[49] Thakur R, Naik A, Panda A et al. Regulation of membrane turnover by phosphatidic acid: cellular functions and disease implications. Frontiers in Cell and Developmental Biology. 2019; 7:83.

[50] Walters DR, Ratsep J, Havis ND. Controlling crop diseases using induced resistance: challenges for the future. J Exp Bot. 2013; 64:1263-80.

[51] Sandroni M, Liljeroth E, Mulugeta T et al. Plant resistance inducers (PRIs): perspectives for future disease management in the field. CABI Reviews. 2020; 1-10:1-10.

[52] Walters DR, Havis ND, Paterson L et al. Cultivar effects on the expression of induced resistance in spring barley. Plant Dis. 2011; 95:595-600.

[53] Sharma K, Butz A, Finckh M. Effects of host and pathogen genotypes on inducibility of resistance in tomato (Solanum lycopersicum) to Phytophthora infestans. Plant Pathol. 2010; 59:1062-71.

[54] Dewen Q, Yijie D, Yi Z et al. Plant immunity inducer development and application. Mol Plant-Microbe Interact. 2017; 30:355-60.

[55] Friedrich L, Lawton K, Dietrich R et al. NIM1 overexpression in Arabidopsis potentiates plant disease resistance and results in enhanced effectiveness of fungicides. Mol Plant-Microbe Interact. 2001; 14:1114-24.

[56] Yu J, Wu S, Sun H et al. CuGenDBv2:an updated database for cucurbit genomics. Nucleic Acids Res. 2023; 51:D1457-64.

[57] Dobin A, Davis CA, Schlesinger F et al. STAR: ultrafast universal RNA-seq aligner. Bioinformatics. 2013; 29:15-21.

[58] Love MI, Huber W, Anders S. Moderated estimation of fold change and dispersion for RNA-seq data with DESeq2. Genome Biol. 2014; 15:1-21.

[59] Bowler C, Benvenuto G, Laflamme P et al. Chromatin techniques for plant cells. Plant J. 2004; 39:776-89.

[60] Nikolaivits E, Agrafiotis A, Termentzi A et al. Unraveling the detoxification mechanism of 2, 4-dichlorophenol by marine-derived mesophotic symbiotic fungi isolated from marine invertebrates. Marine drugs. 2019; 17:564.