Metabolome and Transcriptome Profiling Unveil the Mechanisms of Polyphenol Synthesis in the Developing Endopleura of Walnut (Juglans regia L.)

Abstract

Walnut (Juglans regia L.) is an important woody nut tree species, and its endopleura (the inner coating of a seed) is rich in many polyphenols. Thus far, the pathways and essential genes involved in polyphenol biosynthesis in developing walnut endopleura remain largely unclear. We compared metabolite differences between endopleura and embryo in mature walnuts, and analyzed the changes of metabolites in endopleura at 35, 63, 91, 119, and 147 days after pollination (DAP). A total of 760 metabolites were detected in the metabolome, and the polyphenol contents in endopleura were higher than those in embryos. A total of 15 types of procyanidins, 10 types of kaempferol glycosides, and 21 types of quercetin glycosides that accumulated during endopleura development were identified. The analysis of the phenylpropane metabolic pathway showed that phenylalanine was gradually transformed into proanthocyanidins and other secondary metabolites with the development of endopleura. A total of 49 unigenes related to polyphenol synthesis were identified by transcriptome analysis of endopleura. The expression patterns of PAL, C4H, 4CL, CHS, CHI, F3H, LDOX, and ANR were similar, and their expression levels were highest in endopleura at maturity. Transcriptome and metabolome analysis showed that endopleura rapidly synthesized and accumulated polyphenols during maturation. Moreover, the transcription factor MYB111 played an important role in synthesizing polyphenols in endopleura, and its expression pattern was positively correlated with the accumulation pattern of quercetin, kaempferol, and proanthocyanidins. MYB111 was co-expressed with NAP, NAC, ATR1, and other genes related to cell senescence and abiotic stress response. Our study analyzed the composition and molecular synthesis mechanism of polyphenols in walnut endopleura, and provided new perspectives and insights regarding the nutritional research of walnut nuts.

Integrated metabolomic and transcriptomic dynamic profiles of endopleura coloration during fruit maturation in three walnut cultivars

“…In the grape berry pericarp, UGT88A1 is a key structural gene that is negatively correlated with some flavonoids. The PAL , C4H , 4CL , CHS , CHI , F3H , LDOX , and ANR were highly expressed in the maturity development stage in yellow walnut endopleura [ 33 ]. Most of the CHS (Jr01G10656, Jr02G10304), F3’H (Jr07G12902) and C4H (Jr13G11700) genes involved in polyphenol synthesis are highly expressed during the walnut ripening stage(p5).…”

“…Although the walnut endopleura has a weaker physical protective barrier function compared to the outer shell, it undeniably has a strong chemical defensive function that protects the walnut kernel. This protective function mainly arises from a high concentration of diverse phytochemicals [ 20 , 33 ]. Walnuts contain a significant amount of lipoprotein-bound antioxidants.…”

“…Phenylpropane biosynthesis forms the foundation of the secondary metabolism of polyphenols and flavonoids [ 33 , 63 , 74 ]. Its phenolic intermediates and key enzymes serve to determine the direction of the branching metabolism [ 75 ].…”

“…The walnut endopleura develops from the integument and is generally light yellow, there are specific varieties in which it is purple or bright red [ 20 , 32 ]. Color change of the walnut endopleura are due to polyphenol oxidation and degradation [ 33 ].…”

“…Many beneficial acids, tannins and flavonoids are present in the pericarp or endopleura [ 14 , 33 , 39 ]. There is currently a lack of information on which metabolite changes during the development of purple and red skinned walnuts leads to changes in endopleura color.…”