By performing molecular studies coupled to radiation experiments and in silico systems analyses, we have ascertained the role of the grapevine UV-B receptor and two HY5 homologues in regulating flavonol synthesis.
Stilbene synthase (STS) is the key enzyme leading to the biosynthesis of resveratrol. Recently we reported two R2R3-MYB transcription factor (TF) genes that regulate the stilbene biosynthetic pathway in grapevine: VviMYB14 and VviMYB15. These genes are strongly co-expressed with STS genes under a range of stress and developmental conditions, in agreement with the specific activation of STS promoters by these TFs. Genome-wide gene co-expression analysis using two separate transcriptome compendia based on microarray and RNA sequencing data revealed that WRKY TFs were the top TF family correlated with STS genes. On the basis of correlation frequency, four WRKY genes, namely VviWRKY03, VviWRKY24, VviWRKY43 and VviWRKY53, were further shortlisted and functionally validated. Expression analyses under both unstressed and stressed conditions, together with promoter-luciferase reporter assays, suggested different hierarchies for these TFs in the regulation of the stilbene biosynthetic pathway. In particular, VviWRKY24 seems to act as a singular effector in the activation of the VviSTS29 promoter, while VviWRKY03 acts through a combinatorial effect with VviMYB14, suggesting that these two regulators may interact at the protein level as previously reported in other species.
Background
European grapevine cultivars (
Vitis vinifera spp.) are
highly susceptible to the downy mildew pathogen
Plasmopara viticola
. Breeding of resistant
V. vinifera
cultivars is a promising strategy to reduce the impact of disease management. Most cultivars that have been bred for resistance to downy mildew, rely on resistance mediated by the
Rpv3
(
R
esistance to
P
.
v
iticola
) locus. However, despite the extensive use of this locus, little is known about the mechanism of
Rpv3
-mediated resistance.
Results
In this study,
Rpv3
-mediated defense responses were investigated in
Rpv3
+
and
Rpv3ˉ
grapevine cultivars following inoculation with two distinct
P. viticola
isolates
avrRpv3
+
and
avrRpv3ˉ
, with the latter being able to overcome
Rpv3
resistance. Based on comparative microscopic, metabolomic and transcriptomic analyses, our results show that the
Rpv3–1
-mediated resistance is associated with a defense mechanism that triggers synthesis of fungi-toxic stilbenes and programmed cell death (PCD), resulting in reduced but not suppressed pathogen growth and development. Functional annotation of the encoded protein sequence of genes significantly upregulated during the
Rpv3–1
-mediated defense response revealed putative roles in pathogen recognition, signal transduction and defense responses.
Conclusion
This study used histochemical, transcriptomic and metabolomic analyses of
Rpv3
+
and susceptible cultivars inoculated with avirulent and virulent
P. viticola
isolates to investigate mechanism underlying the
Rpv3–1
-mediated resistance response. We demonstrated a strong correlation between the expressions of stilbene biosynthesis related genes, the accumulation of fungi-toxic stilbenes, pathogen growth inhibition and PCD.
Electronic supplementary material
The online version of this article (10.1186/s12870-019-1935-3) contains supplementary material, which is available to authorized users.
Upon continuous stress exposure, plants display attenuated metabolic stress responses due to regulatory feedback loops. Here, we have tested the hypothesis that pulsed stress exposure with intervening recovery periods should affect these feedback loops, thereby causing increased accumulation of stress-induced metabolites. The response of Arabidopsis plantlets to continuous UV-B exposure (C ) was compared with that of pulsed UV-B exposure (P ). The differential responses to P versus C were monitored at the level of gene expression and metabolite accumulation, using wild type (WT) and different mutant lines. In comparison with C , P increased sinapyl and flavonol (S + F) content, whereas adaptive growth attenuation was reduced. Furthermore, in a myb4 mutant (AtMYB4, repressor-type R2R3-MYB transcription factor), the S + F content was increased only for C , but not beyond the level for P observed in WT. These observations and the ability of AtMYB4 to repress AtMYB12/AtMYB111-mediated activation of target gene promoters (pCHS and pFLS) indicate that the increase of S + F content after P observed in WT plants results from reduced feedback inhibition by AtMYB4. The results support the notion that stress-induced metabolic changes not necessarily cause a growth penalty. Furthermore, the observed P -induced increase in flavonol accumulation may stimulate reevaluation of commercial plant production practices.
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