l-Tryptophan (Trp) is a substrate for the biosynthesis of melatonin, and melatonin is a signal molecule that alters the secondary metabolite profile; whether Trp treatments promote the biosynthesis of melatonin to regulate aroma compounds in grape berries and wine remains unclear. Here, the content of melatonin was higher in Marselan grapes and wine than that in other grape cultivars. Marselan grapes were subjected to 30 different Trp treatments. The content of melatonin and aroma compounds in Marselan grapes was increased by several treatments. Increases in the content of volatiles were mainly driven by increases in aldehydes. Hexanal, the most abundant aldehyde compound, was the main contributor to increases in aldehydes following Trp treatment. The results of our study indicate that the root application of 250 mg/L l-tryptophan during the fruiting expansion stage and the spraying application of 50 mg/L l-tryptophan spray during the veraison stage were the optimal treatments because the content of melatonin and aroma compounds, as well as other basic quality parameters, were highest in Trp-treated grape berries and wine in these treatments. Overall, these two effective Trp treatments could be used to enhance the content of melatonin and aroma compounds in Marselan grapes, and this could increase the economic value of this cultivar.

Melatonin’s role in regulating the biosynthesis of phenolic compounds, particularly stilbenes, remains unknown in grape seeds. In this study, widely targeted metabolomics analysis revealed that 82 phenolic compounds were differentially accumulated in grape seeds from the berries treated with melatonin. The accumulation of resveratrol and its 11 derivatives was largely increased in melatonin-treated berry seeds. Additionally, melatonin treatment of preveraison grape berries increased the resveratrol content in seeds during berry ripening. The results of RNA-Seq showed that a total of 399 genes were differentially expressed, which were mainly involved in the plant hormone signal transduction, metabolic, and biosynthesis of secondary metabolites pathways, in melatonin-treated berry seeds compared to control seeds. Additionally, five PALs, four 4CLs, one C4H, and four STSs were elucidated to be induced by melatonin in seeds at different time points after treatment. Therefore, melatonin promotes resveratrol accumulation and its derivatives, probably by upregulating the genes in the phenylpropanoid pathway, which provides precursors for resveratrol biosynthesis, and the STS genes in grape seeds.

This study determined the composition and content of anthocyanin compounds in red-fleshed 'Kanghong' (KH) berries and wine and revealed the anthocyanin biosynthesis pathway in KH flesh. The 41 anthocyanin compounds detected primarily accumulated in KH skin, followed by flesh and seeds. Anthocyanin compounds with a single glucoside, particularly malvidin-3-O-glucoside, were the most abundant components in KH berries. The percentage of anthocyanin compounds was altered in KH flesh compared to KH skin. KH berries and their wine contained a much higher anthocyanin content than Cabernet Sauvignon (CS) wine. Additionally, a total of 48 differentially accumulated nonanthocyanin phenolic compounds and 2104 differentially expressed genes between KH and CS flesh were identified, and their association analysis indicated that the metabolic flux of phenolic compounds in the phenylpropanoid pathway was promoted in KH flesh. Therefore, this work elucidated the accumulation characteristics of anthocyanins in KH berries and provided the mechanism underlying grape flesh coloration.

This work demonstrated that melatonin increases continuously in seeds, particularly seed coats, during berry ripening. Exogenous melatonin treatments significantly increased the proanthocyanidin (PA) content, partially through ethylene signaling, in seed coats. VvMYB14 expression exhibited patterns similar to melatonin accumulation over time, which was largely induced by melatonin treatment in seed coats during berry ripening. Additionally, VvMYB14 bound to the MBS element of the VvMYBPA1 promoter to activate expression. VvMYB14 overexpression largely upregulated expression of VvMYBPA1, VvMYBPA2 and VvLAR1 and increased the PA content in grape seed-derived calli. Similar increases in AtTT2 and AtBAN expression and PA content were found in VvMYB14-overexpressing Arabidopsis seeds. It was also observed that VvMYB14 overexpression increased ethylene production and thereby induced expression of VvERF104, which bound to the ERF element of the VvMYBPA2 promoter and activated its expression. Additionally, VvERF104 suppression reduced the VvMYB14 overexpression-induced increases in expression of VvMYBPA2 and VvLAR1 and PA content. Further experiments revealed that melatonin-induced increases in the expression of VvMYBPA1, VvMYBPA2, VvERF104 and VvLAR1 and PA accumulation were significantly reduced in VvMYB14-suppressing grape calli and leaves. Collectively, VvMYB14 mediates melatonin-induced PA biosynthesis by directly transactivating VvMYBPA1 expression and indirectly upregulating VvMYBPA2 expression via VvERF104.

Seed development includes an early stage of endosperm proliferation and a late stage of embryo growth at the expense of the endosperm in Arabidopsis thaliana. Abscisic acid (ABA) has known functions during late seed development, but its roles in early seed development remain elusive. In this study, we report that ABA-deficient mutants produced seeds with increased size, mass, and embryo cell number but delayed endosperm cellularization. ABSCISIC ACID DEFICIENT2 (ABA2) encodes a unique short-chain dehydrogenase/reductase that functions in ABA biosynthesis, and its expression pattern overlaps that of SHORT HYPOCOTYL UNDER BLUE1 (SHB1) during seed development. SHB1 RNA accumulation was significantly upregulated in the aba2-1 mutant and was downregulated by the application of exogenous ABA. Furthermore, RNA accumulation of the basic/region leucine zipper transcription factor ABSCISIC ACID-INSENSITIVE5 (ABI5), involved in ABA signaling, was decreased in aba2-1. Consistent with this, seed size was also increased in abi5. We further show that ABI5 directly binds to two discrete regions in the SHB1 promoter. Our results suggest that ABA negatively regulates SHB1 expression, at least in part, through the action of its downstream signaling component ABI5. Our findings provide insights into the molecular mechanisms by which ABA regulates early seed development.