The regulation of the flavonoid pathway has been extensively studied in many plant species. It occurs mostly via the control of the coordinate expression of structural biosynthetic genes by a transcription complex composed of a WD40, a bHLH and a MYB protein. This mechanism has been deeply explored in petunia. In Vitis vinifera, a small set of MYB transcription factors have been characterized and their ability to regulate the synthesis of different flavonoid end-products like anthocyanins or proanthocyanidins have been recently demonstrated. We used functional complementation analyses of well characterized petunia pH/anthocyanin regulatory mutants to gain information about the role of VvMYB5a, VvMYB5b and VvMYBA1 in the regulatory network operating in Vitis vinifera. The coding sequence of VvMYB5a, VvMYB5b and VvMYBA1 was fused to the constitutive promoter 35S and transformed into ph4 and an2 petunia mutant lines. A deep characterization of pigment profile, vacuolar pH and expression of structural genes showed that the restored phenotypes of transformants were attributable to an activation of target genes belonging to the vacuolar acidification or to the anthocyanin pathways. To gain further information about the function of these grape MYB regulators, stable transformation of Vitis vinifera cv Shiraz has been performed to simultaneously silence VvMYB5a and VvMYB5b and, in separate experiments, to over-express each gene independently. All transgenic plants showed apparent abnormal phenotypes suggesting that, unlike the petunia orthologue PhPH4, these regulators may have an important role during the vegetative development in grapevine
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