The Paeonia qiui R2R3-MYB Transcription Factor PqMYB113 Positively Regulates Anthocyanin Accumulation in Arabidopsis thaliana and Tobacco

Paeonia qiui is a wild species of tree peony native to China. Its leaves are purplish red from the bud germination to the flowering stage, and anthocyanin is the main pigment in purplish red leaves. However, the anthocyanin synthesis regulation mechanism in tree peony leaves remains unclear. In this study, an R2R3-MYB, PqMYB113 was identified from the leaves of P. qiui. Phylogenetic analysis revealed that PqMYB113 clustered with Liquidambar LfMYB113 and grape VvMYBA6. Subcellular location analysis showed that PqMYB113 was located in the cell nucleus. The transient reporter assay suggested that PqMYB113 was a transcriptional activator. The overexpression of PqMYB113 in Arabidopsis thaliana and tobacco (Nicotiana tabacum) resulted in increased anthocyanin accumulation and the upregulation of CHS, F3H, F3’H, DFR, and ANS. The dual luciferase reporter assay showed that PqMYB113 could activate the promoters of PqDFR and PqANS. Bimolecular fluorescence complementation assays and yeast two-hybrid assays suggested that PqMYB113 could form a ternary MBW complex with PqbHLH1 and PqWD40 cofactors. These results provide insight into the regulation of anthocyanin biosynthesis in tree peony leaves.

Efficient plant regeneration via meristematic nodule culture in Paeonia ostii ‘Feng Dan’

Tree peony (Paeonia sect. Moutan) is an economically important multipurpose woody plant in terms of its medical, ornamental and oil values, but its breeding and industrial development are severely limited due to inefficient traditional propagation methods and existing in vitro regeneration systems. Meristematic nodules (MNs) are an attractive alternative to solve this problem. This study first presented a protocol for in vitro regeneration of P. ostii ‘Feng Dan’ via MN culture with four consecutive steps, including embryogenic callus (EC) formation, MN induction and leaf cluster differentiation, shoot elongation, rooting and acclimatization. The highest EC induction rate (81.25%) was achieved when cotyledons were cultured on modified Murashige and Skoog (mMS) medium with 4.04 µM N-(2-chloro-4-pyridyl)-N-phenylurea (CPPU) + 5.37 µM α-naphthylacetic acid (NAA) for 30 days. The optimal MN induction rate (100%) and leaf cluster differentiation rate (45.83%) were obtained when ECs were cultured on modified woody plant medium (mWPM) supplemented with 2.02 µM CPPU + 2.27 µM thidiazuron (TDZ) for a subculture time of 10 days. The combination of 1.29 µM 6-benzyladenine (BA) + 0.58 µM gibberellin (GA3) yielded the best shoot elongation (13.40 shoots per nodule), rooting rate (43.33%) and consequently survival rate (45.83%). The study will be beneficial to the mass propagation, breeding and genetic improvement of tree peony.

Genome-Wide Identification and Comparative Profiling of MicroRNAs Reveal Flavonoid Biosynthesis in Two Contrasting Flower Color Cultivars of Tree Peony

MicroRNA (miRNA)-mediated gene regulation is involved in various physiological processes in plants. Flower color is one of the vital ornamental traits of tree peony (Paeonia suffruticosa Andr.). However, the yellow-flowered tree peony cultivars are particularly rare. To elucidate the miRNA-mediated gene regulatory mechanism underlying yellow pigmentation in tree peony, we combined pigment assessment, miRNA identification, expression analysis, and gene functional verification in two contrasting flower color cultivars “High Noon” and “Roufurong.” Flavones/flavonols and anthocyanins were found to be the main contributors to the coloration of “High Noon” and “Roufurong” petals, respectively. Subsequently, miRNA analysis based on available genome data identified 9 differentially expressed miRNAs and 12 relevant target genes implicated in flavonoid biosynthesis. Their dynamic expression patterns determined the key role of mdm-miR156b-PsSPL2 module in yellow pigmentation of tree peony flowers. The sequence analysis and subcellular localization validated that PsSPL2 might function as a nuclear-localized transcription factor. Overexpression of PsSPL2 in tobacco resulted in a decrease of anthocyanin content and down-regulation of NtF3′H and NtDFR transcripts. PsSPL2-silenced petals exhibited lighter yellow color, and the contents of THC, Ap, and Ch decreased significantly. Meanwhile, expression levels of PsCHS, PsCHI, and PsF3H were significantly decreased in the petals with PsSPL2 silencing, while those of PsF3′H and PsDFR were remarkably increased. This study offers a novel insight into yellow pigmentation-related miRNA regulation network in tree peony, and further provides the valuable information on physiological changes during yellow coloring process of tree peony.

Quantitative analysis of resveratrol derivatives in seed coats of tree peonies and their hypoglycemic activities in vitro/vivo

The tree peonies were well-known horticultural and medicinal plants. The tree peony seeds, as emerging woody oil crops, recently have been attracted great attention for their metabolites and bioactivities. In...

A Tree Peony Trihelix Transcription Factor PrASIL1 Represses Seed Oil Accumulation

In many higher plants, seed oil accumulation is governed by complex multilevel regulatory networks including transcriptional regulation, which primarily affects fatty acid biosynthesis. Tree peony (Paeonia rockii), a perennial deciduous shrub endemic to China is notable for its seed oil that is abundant in unsaturated fatty acids. We discovered that a tree peony trihelix transcription factor, PrASIL1, localized in the nucleus, is expressed predominantly in developing seeds during maturation. Ectopic overexpression of PrASIL1 in Nicotiana benthamiana leaf tissue and Arabidopsis thaliana seeds significantly reduced total fatty acids and altered the fatty acid composition. These changes were in turn associated with the decreased expression of multitudinous genes involved in plastidial fatty acid synthesis and oil accumulation. Thus, we inferred that PrASIL1 is a critical transcription factor that represses oil accumulation by down-regulating numerous key genes during seed oil biosynthesis. In contrary, up-regulation of oil biosynthesis genes and a significant increase in total lipids and several major fatty acids were observed in PrASIL1-silenced tree peony leaves. Together, these results provide insights into the role of trihelix transcription factor PrASIL1 in controlling seed oil accumulation. PrASIL1 can be targeted potentially for oil enhancement in tree peony and other crops through gene manipulation.

Effects of Radio Frequency Pretreatment on Quality of Tree Peony Seed Oils: Process Optimization and Comparison with Microwave and Roasting

In this study, we explored the technical parameters of tree peony seeds oil (TPSO) after their treatment with radio frequency (RF) at 0 °C–140 °C, and compared the results with microwave (MW) and roasted (RT) pretreatment in terms of their physicochemical properties, bioactivity (fatty acid tocopherols and phytosterols), volatile compounds and antioxidant activity of TPSO. RF (140 °C) pretreatment can effectively destroy the cell structure, substantially increasing oil yield by 15.23%. Tocopherols and phytosterols were enhanced in oil to 51.45 mg/kg and 341.35 mg/kg, respectively. In addition, antioxidant activities for 2,2-diphenyl-1-picrylhydrazyl (DPPH) and ferric-reducing antioxidant power (FRAP) were significantly improved by 33.26 μmol TE/100 g and 65.84 μmol TE/100 g, respectively (p < 0.05). The induction period (IP) value increased by 4.04 times. These results are similar to those of the MW pretreatment. The contents of aromatic compounds were significantly increased, resulting in improved flavors and aromas (roasted, nutty), by RF, MW and RT pretreatments. The three pretreatments significantly enhanced the antioxidant capacities and oxidative stabilities (p < 0.05). The current findings reveal RF to be a potential pretreatment for application in the industrial production of TPSO.

Integrating full-length transcriptomics and metabolomics reveals the regulatory mechanisms underlying yellow pigmentation in tree peony (Paeonia suffruticosa Andr.) flowers

Tree peony (Paeonia suffruticosa Andr.) is a popular ornamental plant in China due to its showy and colorful flowers. However, yellow-colored flowers are rare in both wild species and domesticated cultivars. The molecular mechanisms underlying yellow pigmentation remain poorly understood. Here, petal tissues of two tree peony cultivars, “High Noon” (yellow flowers) and “Roufurong” (purple–red flowers), were sampled at five developmental stages (S1–S5) from early flower buds to full blooms. Five petal color indices (brightness, redness, yellowness, chroma, and hue angle) and the contents of ten different flavonoids were determined. Compared to “Roufurong,” which accumulated abundant anthocyanins at S3–S5, the yellow-colored “High Noon” displayed relatively higher contents of tetrahydroxychalcone (THC), flavones, and flavonols but no anthocyanin production. The contents of THC, flavones, and flavonols in “High Noon” peaked at S3 and dropped gradually as the flower bloomed, consistent with the color index patterns. Furthermore, RNA-seq analyses at S3 showed that structural genes such as PsC4Hs, PsDFRs, and PsUFGTs in the flavonoid biosynthesis pathway were downregulated in “High Noon,” whereas most PsFLSs, PsF3Hs, and PsF3’Hs were upregulated. Five transcription factor (TF) genes related to flavonoid biosynthesis were also upregulated in “High Noon.” One of these TFs, PsMYB111, was overexpressed in tobacco, which led to increased flavonols but decreased anthocyanins. Dual-luciferase assays further confirmed that PsMYB111 upregulated PsFLS. These results improve our understanding of yellow pigmentation in tree peony and provide a guide for future molecular-assisted breeding experiments in tree peony with novel flower colors.