Lignin is a significant secondary metabolite produced through the phenylpropanoid pathway. As a vital component of the plant cell wall, lignin affects various fruit characteristics, including size, seed quantity, and firmness. In this study, we conducted comprehensive identification and phylogenetic analysis of 265 Caffeic acid O-methyltransferase (COMT) genes across ten different plant species, including Vaccinium corymbosum and four other Vaccinium species. The results reveal that VcCOMT38 is a promising structural gene for the biosynthesis of lignin in blueberry. An in vitro enzymatic assay of VcCOMT38 demonstrated that it is a special enzyme in the lignin biosynthesis pathway and prefers to use caffeic acid as a substrate over 5-hydroxyferulic acid. Transient overexpression and silencing of VcCOMT38 in Vaccinium corymbosum ‘Northland’ fruits demonstrated that VcCOMT38 participates in lignin biosynthesis and contributes to both an increased number of immature seeds and enhanced fruit firmness. The heterologous overexpression of VcCOMT38 in Nicotiana benthamiana revealed that this gene could increase the lignin content and the syringyl/guaiacyl (S/G) ratio, which determines the maximum monomer yield during lignin depolymerization. These results highlight VcCOMT38 as a crucial gene in lignin biosynthesis and its potential for improving lignin production in industry through genetically modified woody plants.

Lotus (Nelumbo spp.) is a valuable plant resource with promising applications in the cosmetic and food industries. To effectively utilize the aromatic compounds of lotus flowers, it is essential to clarify the optimal harvesting organ and flowering stage. In this study, headspace solid-phase microextraction (HS-SPME) and gas chromatography-mass spectrometry (GC—MS) were employed to investigate the volatile profiles of four lotus cultivars across different flowering stages and floral organs. A total of 63 volatile compounds were identified, including 36 terpenes, 10 benzenoids/phenylpropanoids, and 17 fatty acid derivatives. Among them, 1,4-dimethoxybenzene was the dominant aromatic compound. The volatile profiles of petals differed significantly from those of the other five floral organs. The appendages, a component of the stamens, released the highest concentrations of volatiles, with peak emission at the initial-flowering or full-flowering stages. However, due to pollen dispersal, lotus flowers at full flowering were unsuitable for food or cosmetic applications. Thus, appendages harvested during the initial-flowering stage were the most suitable material for high-quality extraction. This study lays a foundation for the industrial development of lotus flower fragrance.

Petal blotch is a prevalent pigmentation pattern observed in the Xibei tree peony (Paeonia rockii), possessing significant aesthetic value and playing a crucial role in the species' reproduction and fitness. Despite years of research, deciphering the molecular mechanisms underlying blotch formation remains challenging. As is well known, floral pigmentation is frequently associated with the familiar R2R3-MYB transcription factors. The key MYB anthocyanin activators of P. rockii ‘Shu Sheng Peng Mo’ were previously reported in our preceding study. In this study, we identified and characterized three R2R3-MYBs, PrMYBi1, PrMYBi2, and PrMYBi3, which belong to subgroup 4 (SG4) and play repressor roles in anthocyanin biosynthesis. A quantitative real-time PCR (qRT-PCR) assay indicated that the expression of PrMYBi1 and PrMYBi3 gradually increased during flowering development and was substantially up-regulated in non-blotch compared to blotch. Yeast one-hybrid and dual-luciferase assays demonstrated that PrMYBi(1–3) directly target the anthocyanin structural genes and repress their transcription. The genetic transformation of tobacco demonstrated that the overexpression of PrMYBi(1–3) decreased anthocyanin accumulation in flowers, with PrMYBi1 serving as the most effective repressor. Our results revealed that SG4 R2R3-MYBs negatively regulate the anthocyanin pathway in P. rockii conservatively, and we provide the definite members. These findings will advance future research to unravel the mystery of blotch pattern formation.

American lotus (Nelumbo lutea) is one of the two species in Nelumbo and has only yellow flower. Identification of total flavonoids showed wild American lotus contained almost only flavonols with quercetin 3-O-glucuronide to be the dominant pigment. The variation tendency of the total flavonol content was coincident with yellow color variation of petals during flower development. To understand the mechanism of accumulation and constituent of pigments in petals, three pivotal genes, NlFLS1, NlFLS2 and NlFLS3, which were predicted to encode flavonol synthases were isolated and characterized by analyses of basic bioinformatics, temporal and spatial expression patterns and enzymatic activity. Their temporal expression levels showed the same variation tendency, which was also consistent with the development-dependent variation of total flavonol content. Spatial expression patterns indicated the three genes should function in petals. All the three proteins were demonstrated to be bifunctional dioxygenase, possessing both flavonol synthase activity and flavanone 3-hydroxylase activity. Besides, other flavonol biosynthesis related genes were also investigated on their expression levels to give more clues on the mechanism. Substrate preferences of the three FLSs, substrate competitions between the FLSs and other flavonol biosynthesis related enzymes, and the greatly differential expression levels between F3'H (flavonoid 3′-hydroxylase) and F3'5'H (flavonoid 3′,5′-hydroxylase) contributed to the flavonol constituent in the petals of America lotus, namely abundant quercetin-derivatives while very few kaempferol-derivatives and myricetin-derivatives.

Previous studies have shown that high light intensity can induce anthocyanin synthesis (AS) in petunia plants. To identify which kind of light quality plays a role in inducing such metabolic process, and what transcripts participate in controlling it, we carried out whole-transcriptome sequencing and analysis of petunia petals treated with different light-quality conditions. Among the red and white light treatments, a total of 2 205 differentially expressed genes and 15, 22, and 20 differentially expressed circRNAs, miRNAs, and lncRNAs, were identified respectively. The AS-related genes, including the structural genes CHSj, F3′H, F3′5′H, DFR, and ANS, and the regulatory genes AN4, DPL, PHZ and MYBx were found to be downregulated under red light condition compared with their levels under white light condition. Furthermore, the light photoreceptor Cryptochrome 3 (CRY3) and a series of light-dependent genes, such as PIF, HY5, and BBXs, were also determined to respond to the light treatments. The anthocyanin contents in early petunia petals under red light were significantly lower than that under white and blue light. The results of qRT-PCR further confirmed the expression pattern of some AS-related and light-response genes in response to different light quality. Yeast two-hybrid results showed that the key elements in the light signal pathway, HY5 can interact with BBX19, BBX24 and BBX25. And PHZ, the important AS regulator can induce anthocyanin synthesis in response to blue light quality from transient expression analysis in petunia petals. These findings presented here not only deepen our understanding of how light quality controls anthocyanin synthesis, but also allow us to explore potential target genes for improving pigment production in petunia flower petals.

Paeonia species are important ornamental plants, including three types: tree peony, herbaceous peony, and Itoh hybrid peony. Paeonia with yellow color is desired and becoming popular. However, the chemical mechanism of the formation of yellow flowers in Paeonia is still unclear. In this study, 14 representative samples were selected: four samples of three species of Paeonia, four herbaceous peony cultivars, three lutea hybrid tree peony cultivars, and three Itoh hybrid peony cultivars. The petal extracts of the samples were determined by HPLC-DAD and HPLC-Q-TOF-MS/MS. In total, 29 flavonoids were isolated and identified, including 28 flavonol glycosides and one chalcone derivative. There were significant differences in the composition and content of flavonoids in petals of different cultivars (species). The total content of flavonoids was between 19.430 and 143.043 mg • g⁻¹ DW. Flavonol glycosides were detected in all samples. Chalconaringenin 2′-O-glucoside was detected from one herbaceous peony cultivar ‘Golden Wheel’ lutea hybrid tree peony cultivars, and Itoh peony cultivars. The content of chalconaringenin 2′-O-glucoside in the samples was more than 43% of total flavonoids. The pigment type and content of lutea hybrid tree peonies and Itoh hybrid peonies are similar. Total flavonoids, especially quercetin 3-O-galloylglucoside and chalconaringenin 2′-O-glucoside contribute greatly to the formation of yellow flowers of Paeonia. The results of this study provide a comprehensive understanding of the chemical mechanism for yellow flower coloration of Paeonia, and lay the foundation for yellow flower breeding of Paeonia.