Reprogramming of primary metabolism facilitates metabolic engineering of bioactives in tomato fruit (ChIP-Seq). Solanum lycopersicum

Phenylpropanoids are derived from phenylalanine and comprise an important class of plant secondary metabolites that include specialized bioactives with medicinal properties, important phytonutrients, a broad range of natural colours, phytoanticipins, phytoalexins and phytoestrogens. A number of transcription factors have been used to upregulate specific branches of phenylpropanoid metabolism, but by far the most effective has been the fruit-specific expression of AtMYB12 in tomato, which resulted in an astonishing 10% of fruit dry weight accumulating as flavonoids and hydroxycinnamates. We show that AtMYB12 not only increases the demand of flavonoid biosynthesis, but also increases the supply of carbon from primary metabolism, and the supply of energy and reducing power, by upregulating glycolysis, the TCA cycle, the oxidative pentose phosphate pathway, fuelling the shikimate and phenylalanine biosynthetic pathways to supply more aromatic amino acids for secondary metabolism. AtMYB12 directly activates at least some genes encoding enzymes of primary metabolism. The enhanced supply of precursors, energy and reducing power achieved by AtMYB12 expression can be harnessed to engineer high levels of novel phenylpropanoids in tomato fruit, offering an effective production system for bioactives and other high value ingredients. Overall design: ChIP-Seq experiments were done using AtMYB12 tomato fruit at 3-4 days after breaker (when the expression of AtMYB12 is strongly induced).

苯丙素类（Phenylpropanoids）源自苯丙氨酸，是一类重要的植物次生代谢产物，涵盖了具有药用特性的特异性生物活性成分、关键植物营养素、多种天然色素、植物预存抗菌素（phytoanticipins）、植物诱导型抗毒素（phytoalexins）以及植物雌激素（phytoestrogens）。已有多种转录因子被应用于上调苯丙素代谢的特定分支，但目前已知成效最为显著的策略是在番茄中实现AtMYB12的果实特异性表达，该操作可使番茄果实干重中高达10%的组分以类黄酮与羟基肉桂酸类的形式积累。我们的研究表明，AtMYB12不仅提升了类黄酮生物合成的底物需求，还通过上调糖酵解、三羧酸（TCA）循环与氧化戊糖磷酸途径，增加了初生代谢的碳供给以及能量与还原当量的供给，为莽草酸与苯丙氨酸生物合成通路供能，进而为次生代谢提供更多芳香族氨基酸。此外，AtMYB12可直接激活至少部分编码初生代谢相关酶的基因。通过AtMYB12表达所实现的前体、能量与还原当量供给提升，可被用于工程化改造番茄果实以积累高水平的新型苯丙素类物质，为生物活性成分及其他高价值产物提供了高效的生产平台。实验设计概述：以破色期（breaker）3-4天的转AtMYB12基因番茄果实为材料开展染色质免疫共沉淀测序（ChIP-Seq）实验，该时期AtMYB12的表达会被强烈诱导。