Natural Variation in Monoterpene Synthesis in Kiwifruit: Transcriptional Regulation of Terpene Synthases by NAC and ETHYLENE-INSENSITIVE3-Like Transcription Factors

• Published in: Plant physiology (Bethesda) Vol. 167; no. 4; pp. 1243 - 1258
• Main Authors: Nieuwenhuizen, Niels J., Chen, Xiuyin, Wang, Mindy Y., Matich, Adam J., Perez, Ramon Lopez, Allan, Andrew C., Green, Sol A., Atkinson, Ross G.
• Format: Journal Article
• Language: English
• Published: United States American Society of Plant Biologists 01.04.2015
• Subjects: Actinidia - enzymology; Actinidia - genetics; Actinidia - growth & development; Alkyl and Aryl Transferases - genetics; Alkyl and Aryl Transferases - metabolism; Base Sequence; Binding sites; BIOCHEMISTRY AND METABOLISM; Enzymes; Erythritol - analogs & derivatives; Erythritol - metabolism; Ethylenes - metabolism; Fruit - enzymology; Fruit - genetics; Fruit - growth & development; Fruits; Gene Expression; Gene expression regulation; Gene Expression Regulation, Plant; Genes; Molecular Sequence Data; Monoterpenes; Monoterpenes - metabolism; Phylogeny; Plant Leaves - enzymology; Plant Leaves - genetics; Plant Proteins - genetics; Plant Proteins - metabolism; Plants; Plants, Genetically Modified; Promoter Regions, Genetic - genetics; Ripening; Sequence Alignment; Sequence Analysis, DNA; Species Specificity; Sugar Phosphates - metabolism; Terpenes; Transcription factors; Transcription Factors - genetics; Transcription Factors - metabolism; Transferases - genetics; Transferases - metabolism
• DOI: 10.1104/pp.114.254367
• ISSN: 0032-0889; 1532-2548

Two kiwifruit (Actinidia) species with contrasting terpene profiles were compared to understand the regulation of fruit monoterpene production. High rates of terpinolene production in ripeActinidia argutafruit were correlated with increasing gene and protein expression ofA. argutaterpene synthase1 (AaTPS1) and correlated with an increase in transcript levels of the 2-C-methyl-D-erythritol 4-phosphate pathway enzyme 1-deoxy-D-xylulose-5-phosphate synthase (DXS).Actinidia chinensis terpene synthase1(AcTPS1) was identified as part of an array of eight tandemly duplicated genes, andAcTPS1expression and terpene production were observed only at low levels in developing fruit. Transient overexpression ofDXSinNicotiana benthamianaleaves elevated monoterpene synthesis by AaTPS1 more than 100-fold, indicating that DXS is likely to be the key step in regulating 2-C-methyl-D-erythritol 4-phosphate substrate flux in kiwifruit. Comparative promoter analysis identified potential NAC (for no apical meristem [NAM], Arabidopsis transcription activation factor [ATAF], and cup-shaped cotyledon [CUC])-domain transcription factor) and ETHYLENE-INSENSITIVE3-like transcription factor (TF) binding sites in theAaTPS1promoter, and cloned members of both TF classes were able to activate theAaTPS1promoter in transient assays. Electrophoretic mobility shift assays showed that AaNAC2, AaNAC3, and AaNAC4 bind a 28-bp fragment of the proximal NAC binding site in theAaTPS1promoter but not theA. chinensis AcTPS1promoter, where the NAC binding site was mutated. Activation could be restored by reintroducing multiple repeats of the 12-bp NAC core-binding motif. The absence of NAC transcriptional activation in ripeA. chinensisfruit can account for the low accumulation ofAcTPS1transcript, protein, and monoterpene volatiles in this species. These results indicate the importance of NAC TFs in controlling monoterpene production and other traits in ripening fruits.