Author(s): Santos, Catarina Antunes dos
Date: 2023
Persistent ID: http://hdl.handle.net/10400.5/95455
Origin: Repositório da Universidade de Lisboa
Subject(s): Vitis vinifera; Plasmopara viticola; fosfolipase C; fosfolipase D; sinalização lipídica; Teses de mestrado - 2023
Vitis vinifera L. is presently one of the most cultivated fruit crops in the world, being highly susceptible to Plasmopara viticola (Berk. & M.A. Curtis) Berl. & De Toni, which causes the grapevine downy mildew, leading to vast yield reductions and, consequently, to massive economic losses. Therefore, with the known adverse effects of pesticides on the environment and human health, it is crucial the improvement and application of alternative approaches and techniques against this disease. Lipid-signalling enzymes, such as phospholipases, have been reported as essential components of lipid-associated signalling to several specific biotic and abiotic stresses. Although phosphatidic acid has been described as a crucial player on defense mechanisms on several plant species this has not been tested in grapevine. Phosphatidic acid is produced from membrane phospholipids via phospholipases C or as a product of phospholipases D activity. Phospholipases C include phosphoinositide-specific phospholipases C and the nonspecific phospholipases C, whereas phospholipase D family include C2- PLDs, PXPH-PLDs and SP-PLDs. In this work, we were able to identify 11 genes coding for 13 grapevine phospholipase C proteins and 11 genes coding for 15 grapevine phospholipase D proteins. This study provides in silico characterization of grapevine phospholipases C and D gene family members, with the main aim of evaluate their lipid-signalling roles in grapevine - pathogen interactions and immune priming events. Therefore, three V. vinifera cultivars with different resistant degrees to P. viticola were selected, namely: one susceptible genotype “Chardonnay”; one tolerant genotype “Regent” that harbours an Rpv3.1 resistance loci and one resistant genotype “Sauvignac” that harbours a pyramid loci of Rpv12 and Rpv3.1 resistance loci, both mock inoculated and inoculated with NW-10/16, an extremely virulent strain on Rpv3 genotypes. In “Chardonnay” and “Regent” this isolate was able to overcome the hypersensitive response, fully developing and releasing sporangia, whereas in “Sauvignac” it was not capable to overcome its resistance. Alterations of leaf gene expression at different time-points after infection indicate that phospholipases C and D are involved in grapevine defense mechanisms. Furthermore, “Chardonnay” and “Sauvignac” presented similar expression patterns for most of the analysed genes which may suggest that the lipid associated pathway is not required for “Sauvignac” defense response. There were basal differences in fatty acids contents and composition between susceptible and tolerant cultivars and also a different modulation after P. viticola inoculation, which is in agreement with previous results. Moreover, the susceptible cultivar presents more unsaturated fatty acids and higher Double Bound Index, which may suggest that membranes are more fluid and therefore facilitate pathogen entry. Higher levels of total lipids in the inoculated susceptible grapevine genotype are in accordance with lower expression levels found for phospholipases A, C and D associated genes.
