REGULATORY MECHANISMS CONTROLLING FLAVONOID AND VACUOLAR ACIDIFICATION PATHWAYS IN GRAPEVINE

In a wide range of plants, a common regulatory complex composed by a MYB and a bHLH TF and a WD40 regulatory protein initiates multiple cellular differentiation pathways. Although the MYB proteins are the most responsible for complex specificity, other intermediate regulators could be required for the activation of a specific set of genes. In Arabidopsis the WRKY TF TTG2 is under the control of the regulatory complex driven by MYB5 in the regulation of trichome development and proanthocyanidin and mucilage production in seed coat (Johnson et al., 2002; Ishida et al., 2007). In Petunia, another regulator belonging to the WRKY family, PH3, acts down-stream and in concert with the MYB protein PH4-driven complex in the control of vacuolar acidification in petals epidermis (Verweij et al., 2016). Among grapevine MYB and WRKY TFs, MYB5a and MYB5b are both the orthologs to AtMYB5 and PhPH4, and WRKY26 has been identified as the closer homolog to AtTTG2 and PhPH3 (Deluc et al., 2008; Wang et al., 2014, Verweij et al., 2016). We therefore hypothesized that WRKY26 could be an intermediate regulator acting down-stream a regulatory complex involving MYB5a and MYB5b. In this work of thesis, we observed high correlation between WRKY26 and MYB5a expression and by in situ hybridization we demonstrated that they even co-localized in specific cell types in berry tissues. We also demonstrated that WRKY26 promoter region is highly activated by the combined action of MYB5a and WRKY26, suggesting that WRKY26 could acts down-stream MYB5a, but also in concert with the regulatory complex it belongs. Ultimately, testing the ability of the proposed MYB5a/MYB5b-WRKY26 network to activate specific target genes, we demonstrated that WRKY26 could provide more variability to the processes regulated by the aforementioned network. Above all, our results strongly suggest that in grapevine MYB5a/MYB5b and WRKY26 may act in a transcriptional regulatory network likely conserved among plant species and controlling both flavonoid and vacuolar acidification pathways. Flavonoids content and vacuolar acidity play important roles in grapevine in determining the quality of fruits and of wine and further investingation aimed to dissect common regulatory mechanisms controlling both these traits could represent an important advance in the possibility to control the final berry quality traits for winemaking.