Plant secondary metabolism is complex in its diverse chemical composition and dynamic regulation of biosynthesis. How the functional diversification of enzymes contributes to the diversity is largely unknown. In the flavonoids pathway, dihydroflavonol 4-reductase (DFR) is a key enzyme mediating dihydroflavanol into anthocyanins biosynthesis. Here, the DFR homolog was identified from Camellia nitidissima Chi. (CnDFR) which is a unique species of the genus Camellia with golden yellow petals. Sequence analysis showed that CnDFR possessed not only conserved catalytic domains, but also some amino acids peculiar to Camellia species. Gene expression analysis revealed that CnDFR was expressed in all tissues and the expression of CnDFR was positively correlated with polyphenols but negatively with yellow coloration. The subcellular localization of CnDFR by the tobacco infiltration assay showed a likely dual localization in the nucleus and cell membrane. Furthermore, overexpression transgenic lines were generated in tobacco to understand the molecular function of CnDFR. The analyses of metabolites suggested that ectopic expression of CnDFR enhanced the biosynthesis of polyphenols, while no accumulation of anthocyanins was detected. These results indicate a functional diversification of the reductase activities in Camellia plants and provide molecular insights into the regulation of floral color.

Leaf nitrogen (N) and phosphorus (P) stoichiometry at a large geographical scale is the result of long-term adaptation to the environment. Therefore, the patterns of leaf N and P spatial distributions and their controlling factors represent an important issue in current ecological research. To explore the leaf stoichiometry of Chinese fir at a national level, we conducted a meta-analysis based on the dataset of the leaf nitrogen (N) and phosphorus (P) concentrations and the N:P ratio from 28 study sites across China. For all of the age groups considered, the average concentrations of the leaf N and P concentrations and the N:P ratio were 11.94 mg g⁻¹, 1.04 mg g⁻¹, and 12.93, respectively. Significant differences were found in the leaf P concentration and N:P ratio between the five age groups, while the differences in the leaf N concentration between the groups were not significant. Linear fitting results indicated that the leaf P concentration decreased, and the leaf N:P ratio increased with the increase of the MAT (mean annual temperature) and soil N concentration. Redundancy analysis (RDA) revealed that the first axis, with an explanatory quantity of 0.350, indicated that the MAT (mean annual temperature), soil nitrogen concentration and stand age had a good relationship with the leaf P concentration and N:P ratio, while the second axis, with an explanatory quantity of 0.058, indicated that the leaf N concentration was less affected by the environmental factors. These results demonstrate that the leaf P concentration and N:P ratio are affected by the stand age, an uneven distribution of the heat and soil nutrient concentration status, and N, as the limiting element, remaining relatively stable. Overall, our findings revealed the response of leaf stoichiometric traits to environment change, which benefits the management of Chinese fir plantations.

Camellia nitidissima Chi is an ornamental plant of the genus Camellia L. Its flowers contain a lot of flavonoids and polyphenols. Flavonoid 3'-hydroxylase (F3'H) plays an important role in the synthesis of flavonoids, polyphenols and anthocyanins. We cloned CnF3'H from the petal of C. nitidissima (GenBank code: HQ290518.1). The full length of CnF3'H was 1859 bp, with an open reading frame of 1577 bp, and encoded 518 amino acid. A phylogenetic tree analysis showed that CnF3'H was closely related to Camellia sinensis L. and C. sinensis cultivar Zhonghuang. CnF3'H was expressed in flowers, leaves, fruits, sepals, petals and stamens of C. nitidissima, and during the flowering process the expression level in flower decreased initially and then increased. CnF3'H expression was significantly positive correlated with polyphenol contents in C. nitidissima. A CnF3'H-EGFP expression vector was constructed to do the subcellular localization, we found that CnF3'H was obviously localized in the nuclear envelope and cytomembrane. In transgenic tobacco flowers, the total polyphenol content and various polyphenol constituents were significantly increased with high CnF3'H expression level, while total flavonoid contents and some flavonol constituents were increased slightly. These findings suggest that CnF3'H can promote the synthesis of polyphenols better than flavonoids.