ASSOCIATION OF THE EXPRESSION LEVELS OF TRANSCRIPTION FACTORS WITH THE PHENOTYPES AND GENOTYPES OF PEACH FRUITS THAT DIFFER IN THEIR QUALITATIVE CHARACTERISTICS

Anthocyanins are plant bioactive compounds that control color of many vegetables, flowers, fruit, roots and other plant parts. In peach [Prunus persica (L.) Batsch], color is a key determinant for fruit quality and depends by flovonoids including anthocyanins. These bioactive molecules have potential benefits to human health including protection against cancer, cardiovascular diseases, inflammation and other chronic diseases. The R2R3-MYB transcription factors (TFs) control the expression of genes for the synthesis of anthocyanin pigments with the help of co-activators belonging to the basic-helix-loop-helix (bHLH) and WD40 repeat family. In the peach genome, through this study three MYB10-like (MYB10.1, MYB10.2 and MYB10.3) and three bHLH-like (bHLH3, bHLH33 and GL3) TFs were identified as best candidates to be the regulators of the anthocyanin accumulation in peach. Among these MYB10s, MYB10.1 is the highly expressed gene in ripe peach fruits. The accumulation of anthocyanins in yellow flesh peach fruits is highest in the peel, abundant in the part of the mesocarp surrounding the stone and lowest in the mesocarp. The expression of MYB10.1 and MYB10.3 genes were correlated with anthocyanin levels of different peach parts. They have also positive correlation with the key structural genes of the anthocyanin pathway, like chalcone synthase (CHS), flavanone-3?-hydroxylase (F3H), dihydroflavonol 4-reductase (DFR) and UDP-glucose:flavonoid-3-O-glucosyltransferase (UFGT). Functions of peach MYB10s were tested in tobacco and shown to activate key genes in the anthocyanin pathway when bHLHs were co-expressed as partners. Over-expression of MYB10.1/bHLH3 and MYB10.3/bHLH3 activated anthocyanin production by up-regulating NtCHS, NtDFR and NtUFGT while other combinations were not effective. To better understand the role of MYB10.1 in anthocyanin synthesis during peach ripening, its coding sequence has been constitutively expressed in tobacco transgenic plants. Its over-expression induce anthocyanin pigmentation in the reproductive parts of transgenic tobacco lines through the up-regulation of transcript levels of many important genes involved in the anthocyanin biosynthetic pathway like phenylalanineammonia-lyase (NtPAL), NtCHS, chalcone isomerase (NtCHI), NtF3H, NtDFR, anthocyanin synthase (NtANS) and NtUFGT. The pigment accumulation was always limited to reproductive parts and never present in the vegetative part like stem and leaves. Besides the regulation of anthocyanin biosynthesis, the ectopic expression of MYB10.1 leads to two different types of phenotype in transgenic as compared to WT tobacco. The strong phenotype, called type-I, is characterized by irregular leaf shape and size, reduced plant height, stamens with reduced filament length and non-dehiscing anthers, reduced pistil length, no nectary gland formation and reduced capsule development but pigmented reproductive parts including androecium, gynoecium and petals. On the contrary, so-called type-II plants, have a mild phenotype with regular plant growth and development, but purple seed coats. Surprisingly, the over-expression of MYB10.1 altered the floral development in transgenic tobacco lines. In type-I transgenic plants, the over-expression of peach MYB10.1 leads to down-regulation of NtMYB305, a gene similar, but not orthologous, to MYB10.1, which is required for floral development. Moreover, MYB10.1 up-regulates the expression of jasmonic acid (JA) biosynthesis (allene oxide synthase, NtAOS) and signaling (NtJAZd) pathway genes as well as 1-aminocyclopropane-1-carboxylate oxidase (NtACO), a key gene in ethylene synthesis. Furthermore, NECTARIN1 (NtNCE1) that is known to be involved in nectary gland formation, was repressed since its transcription is regulated by NtMYB305. On the contrary, type-II transgenic plants showed opposite results for NtMYB305, NtAOS, NtJAZd and NtNEC1. Therefore, it can be concluded that the over-expression of peach MYB10.1 in tobacco not only regulates flavonoid biosynthesis in the reproductive parts but also affects (directly and/or indirectly) other process like vegetative and reproductive development. These new findings will promote further investigation to elucidate more diverse role of R2R3-MYB TFs. Furthermore, they will promote more and better insights on the regulation of anthocyanin biosynthesis and will aid fruit breeders to introduce new peach cultivars with higher antioxidant level as well as colored fruit.