Dynamic DNA Methylation Regulates Season-Dependent Secondary Metabolism in the New Shoots of Tea Plants.

Plant secondary metabolites are critical quality-conferring compositions of plant-derived beverages, medicines, and industrial materials. The accumulations of secondary metabolites are highly variable among seasons; however, the underlying regulatory mechanism remains unclear, especially in epigenetic regulation. Here, we used tea plants to explore an important epigenetic mark DNA methylation (5mC)-mediated regulation of plant secondary metabolism in different seasons. Multiple omics analyses were performed on spring and summer new shoots. The results showed that flavonoids and theanine metabolism dominated in the metabolic response to seasons in the new shoots. In summer new shoots, the genes encoding DNA methyltransferases and demethylases were up-regulated, and the global CG and CHG methylation reduced and CHH methylation increased. 5mC methylation in promoter and gene body regions influenced the seasonal response of gene expression; the amplitude of 5mC methylation was highly correlated with that of gene transcriptions. These differentially methylated genes included those encoding enzymes and transcription factors which play important roles in flavonoid and theanine metabolic pathways. The regulatory role of 5mC methylation was further verified by applying a DNA methylation inhibitor. These findings highlight that dynamic DNA methylation plays an important role in seasonal-dependent secondary metabolism and provide new insights for improving tea quality.

Molecular and Biochemical Characterization of Jasmonic Acid Carboxyl Methyltransferase Involved in Aroma Compound Production of Methyl Jasmonate during Black Tea Processing.

Methyl jasmonate (MeJA), a volatile organic compound, is a principal flowery aromatic compound in tea. During the processing of black tea, MeJA is produced by jasmonic acid carboxyl methyltransferase (JMT) of the jasmonic acid (JA) substrate, forming a specific floral fragrance. CsJMT was cloned from tea leaves; the three-dimensional structure of CsJMT was predicted. Enzyme activity was identified, and protein purification was investigated. Site-directed deletions revealed that N-10, S-22, and Q-25 residues in the beginning amino acids played a key functional role in enzyme activity. The expression patterns of CsJMT in tea organs differed; the highest expression of CsJMT was observed in the fermentation process of black tea. These results aid in further understanding the synthesis of MeJA during black tea processing, which is crucial for improving black tea quality using specific fragrances and could be applied to the aromatic compound regulation and tea breeding improvement in further studies.

Enantiomeric Trimethylallantoin Monomers, Dimers, and Trimethyltriuret: Evidence for an Alternative Catabolic Pathway of Caffeine in Tea Plant.

Racemic trimethylallantoin monomer (1), mesomeric and racemic trimethylallantoin dimers (2 and 3), were isolated from tea. Two pairs of optically pure enantiomers (1a, 1b and 3a, 3b) were separated by chiral column from the two racemes (1 and 3). Their structures were elucidated by a combination of extensive spectroscopic techniques, single-crystal X-ray diffraction, and experimental and calculated electronic circular dichroism. A novel caffeine catabolic pathway was proposed based on the caffeine stable isotopic tracer experiments.

Caffeine Content and Related Gene Expression: Novel Insight into Caffeine Metabolism in Camellia Plants Containing Low, Normal, and High Caffeine Concentrations.

Caffeine is a crucial secondary metabolic product in tea plants. Although the presence of caffeine in tea plants has been identified, the molecular mechanisms regulating relevant caffeine metabolism remain unclear. For the elucidation of the caffeine biosynthesis and catabolism in Camellia plants, fresh, germinated leaves from four Camellia plants with low (2), normal (1), and high (1) caffeine concentrations, namely, low-caffeine tea 1 (LCT1, Camellia crassicolumna), low-caffeine tea 2 (LCT2, C. crassicolumna), Shuchazao (SCZ, C. sinensis), and Yunkang 43 (YK43, C. sinensis) were used in this research. Transcriptome and purine alkaloids analyses of these Camellia leaves were performed using RNA-Seq and liquid chromatography-mass spectrometry (LC-MS). Moreover, 15N-caffeine tracing was performed to determine the metabolic fate of caffeine in leaves of these plants. Caffeine content was correlated with related gene expression levels, and a quantitative real-time (qRT) PCR analysis of specific genes showed a consistent tendency with the obtained transcriptomic analysis. On the basis of the results of stable isotope-labeled tracer experiments, we discovered a degradation pathway of caffeine to theobromine. These findings could assist researchers in understanding the caffeine-related mechanisms in Camellia plants containing low, normal, and high caffeine content and be applied to caffeine regulation and breeding improvement in future research.

Significantly increased amino acid accumulation in a novel albino branch of the tea plant (Camellia sinensis).

MAIN CONCLUSION: A normal tea plant with one albino branch was discovered. RNA sequencing, albinism phenotype and ultrastructural observations provided a valuable understanding of the albino mechanism in tea plants. Tea plants with a specific color (white or yellow) have been studied extensively. A normal tea plant (Camellia sinensis cv. quntizhong) with one albino branch was discovered in a local tea plantation in Huangshan, Anhui, China. The pure albino leaves on this special branch had accumulated a fairly high content of amino acids, especially theanine (45.31 mg/g DW), and had a low concentration of polyphenols and an extremely low chlorophyll (Chl) content compared with control leaves. Ultrastructural observation of an albino leaf revealed no chloroplasts, whereas it was viable in the control leaf. RNA sequencing and differentially expressed gene (DEG) analysis were performed on the albino leaves and on control leaves from a normal green branch. The related genes involved in theanine and polyphenol biosynthesis were also investigated in this study. DEG expression patterns in Chl biosynthesis or degradation, carotenoid biosynthesis or degradation, chloroplast development, and biosynthesis were influenced in the albino leaves. Chloroplast deletion in albino leaves had probably destroyed the balance of carbon and nitrogen metabolism, leading to a high accumulation of free amino acids and a low concentration of polyphenols in the albino leaves. The obtained results can provide insight into the mechanism underlying this special albino branch phenotype, and are a valuable contribution toward understanding the albino mechanism in tea plants.