Data_Sheet_1_The Carotenoid Esterification Gene BrPYP Controls Pale-Yellow Petal Color in Flowering Chinese Cabbage (Brassica rapa L. subsp. parachinensis).docx

Carotenoid esterification plays indispensable roles in preventing degradation and maintaining the stability of carotenoids. Although the carotenoid biosynthetic pathway has been well characterized, the molecular mechanisms underlying carotenoid esterification, especially in floral organs, remain poorly understood. In this study, we identified a natural mutant flowering Chinese cabbage (Caixin, Brassica rapa L. subsp. chinensis var. parachinensis) with visually distinguishable pale-yellow petals controlled by a single recessive gene. Transmission electron microscopy (TEM) demonstrated that the chromoplasts in the yellow petals were surrounded by more fully developed plastoglobules compared to the pale-yellow mutant. Carotenoid analyses further revealed that, compared to the pale-yellow petals, the yellow petals contained high levels of esterified carotenoids, including lutein caprate, violaxanthin dilaurate, violaxanthin-myristate-laurate, 5,6epoxy-luttein dilaurate, lutein dilaurate, and lutein laurate. Based on bulked segregation analysis and fine mapping, we subsequently identified the critical role of a phytyl ester synthase 2 protein (PALE YELLOW PETAL, BrPYP) in regulating carotenoid pigmentation in flowering Chinese cabbage petals. Compared to the yellow wild-type, a 1,148 bp deletion was identified in the promoter region of BrPYP in the pale-yellow mutant, resulting in down-regulated expression. Transgenic Arabidopsis plants harboring beta-glucuronidase (GUS) driven by yellow (BrPYPY::GUS) and pale-yellow type (BrPYPPY::GUS) promoters were subsequently constructed, revealing stronger expression of BrPYPY::GUS both in the leaves and petals. Furthermore, virus-induced gene silencing of BrPYP significantly altered petal color from yellow to pale yellow. These findings demonstrate the molecular mechanism of carotenoid esterification, suggesting a role of phytyl ester synthase in carotenoid biosynthesis of flowering Chinese cabbage.

类胡萝卜素酯化作用在防止类胡萝卜素降解和维持其稳定性方面发挥着不可或缺的作用。尽管类胡萝卜素生物合成途径已被充分表征，但类胡萝卜素酯化的分子机制，尤其是在花器官中，仍理解不足。在本研究中，我们鉴定了一种自然突变型花椰菜（菜心，Brassica rapa L. subsp. chinensis var. parachinensis），其花瓣呈可观察的淡黄色，由单个隐性基因控制。透射电子显微镜（TEM）显示，与淡黄色突变体相比，黄色花瓣中的油滴被更完全发育的质体小球所包围。类胡萝卜素分析进一步揭示，与淡黄色花瓣相比，黄色花瓣含有高水平的酯化类胡萝卜素，包括叶黄酸硬脂酯、紫黄质二硬脂酯、紫黄质-肉豆蔻酸-硬脂酯、5,6-环氧叶黄酸二硬脂酯、叶黄酸二硬脂酯和叶黄酸硬脂酯。基于成束分离分析和精细定位，我们随后确定了植物醇酯合成酶2蛋白（PALE YELLOW PETAL，BrPYP）在调控花椰菜花瓣类胡萝卜素色素化中的关键作用。与黄色野生型相比，在淡黄色突变体中BrPYP启动子区域存在1,148 bp的缺失，导致表达下调。随后构建了携带黄色（BrPYPY::GUS）和淡黄色型（BrPYPPY::GUS）启动子的转基因拟南芥植物，结果表明BrPYPY::GUS在叶片和花瓣中的表达更强。此外，BrPYP的病毒诱导基因沉默显著改变了花瓣颜色，从黄色变为淡黄色。这些发现揭示了类胡萝卜素酯化的分子机制，表明植物醇酯合成酶在花椰菜花瓣类胡萝卜素生物合成中发挥作用。