Background: Wintersweet (Chimonanthus praecox), an important ornamental plant, has evolved unique fragrant aroma and winter-flowering properties, which are critical for its successful sexual reproduction. However, the molecular mechanisms underlying these traits are largely unknown in this species. In addition, wintersweet is also a typical representative species of the magnoliids, where the phylogenetic position of which relative to eudicots and monocots has not been conclusively resolved. Results: Here, we present a chromosome-level wintersweet genome assembly with a total size of 695.36 Mb and a draft genome assembly of Calycanthus chinensis. Phylogenetic analyses of 17 representative angiosperm genomes suggest that Magnoliids and eudicots are sister to monocots. Whole-genome duplication signatures reveal two major duplication events in the evolutionary history of the wintersweet genome, with an ancient one shared by Laurales, and a more recent one shared by the Calycantaceae. Whole-genome duplication and tandem duplication events have significant impacts on copy numbers of genes related to terpene and benzenoid/phenylpropanoid (the main floral scent volatiles) biosynthesis, which may contribute to the characteristic aroma formation. An integrative analysis combining cytology with genomic and transcriptomic data reveals biological characteristics of wintersweet, such as floral transition in spring, floral organ specification, low temperature-mediated floral bud break, early blooming in winter, and strong cold tolerance. Conclusions: These findings provide insights into the evolutionary history of wintersweet and the relationships among the Magnoliids, monocots, and eudicots; the molecular basis underlying floral scent biosynthesis; and winter flowering, and highlight the utility of multi-omics data in deciphering important ornamental traits in wintersweet.

Wintersweet (Chimonanthus praecox L.) is an ornamental and economically significant shrub known for its unique flowering characteristics, especially the emission of abundant floral volatile organic compounds. Thus, an understanding of the molecular mechanism of the production of these compounds is necessary to create new breeds with high volatile production. In this study, two bHLH transcription factors (CpMYC2 and CpbHLH13) of Wintersweet H29 were functionally characterized to illustrate their possible role in the production of volatile compounds. The qRT-PCR results showed that the expression of CpMYC2 and CpbHLH13 increased from the flower budding to full bloom stage, indicating that these two genes may play an essential role in blooming and aroma production in wintersweet. Gas chromatography-mass spectroscopy (GC-MS) analysis revealed that the overexpression of CpMYC2 in arabidopsis (Arabidopsis thaliana) AtMYC2-2 mutant (Salk_083483) and tobacco (Nicotiana tabaccum) genotype Petit Havana SR1 significantly increased floral volatile monoterpene, especially linalool, while the overexpression of CpbHLH13 in Arabidopsis thaliana ecotype Columbia-0 (Col-0) and tobacco genotype SR1 increased floral sesquiterpene β-caryophyllene production in both types of transgenic plants respectively. High expression of terpene synthase (TPS) genes in transgenic A. thaliana along with high expression of CpMYC2 and CpbHLH13 in transgenic plants was also observed. The application of a combination of methyl jasmonic acid (MeJA) and gibberellic acid (GA3) showed an increment in linalool production in CpMYC2-overexpressing arabidopsis plants, and the high transcript level of TPS genes also suggested the involvement of CpMYC2 in the jasmonic acid (JA) signaling pathway. These results indicate that both the CpMYC2 and CpbHLH13 transcription factors of wintersweet are possibly involved in the positive regulation and biosynthesis of monoterpene (linalool) and sesquiterpene (β-caryophyllene) in transgenic plants. This study also indicates the potential application of wintersweet as a valuable genomic material for the genetic modification of floral scent in other flowering plants that produce less volatile compounds.

Anthocyanins play a paramount role in color variation and significantly contribute to the economic value of ornamental plants. The conserved activation complex MYB-bHLH-WD40 (MBW; MYB: v-myb avian myeloblastosis viral oncogene homolog; bHLH: basic helix–loop–helix protein; WD40:WD-repeat protein) involved in anthocyanin biosynthesis has been thoroughly researched, but there have been limited investigations into the function of repressor factors. In this study, we characterized TgMYB4, an R2R3-MYB transcriptional repressor which is highly expressed during petal coloration in red petal cultivars. TgMYB4-overexpressing tobaccos exhibited white or light pink petals with less anthocyanin accumulation compared to control plants. TgMYB4 was found to inhibit the transcription of ANTHOCYANIDIN SYNTHASE (TfANS1) and DIHYDRO-FLAVONOL-4-REDUCTASE (AtDFR), although it did not bind to their promoters. Moreover, the TgMYB4 protein was able to compete with the MYB activator to bind to the :bHLHprotein, thereby suppressing the function of the activator MBW complex. These findings demonstrate that TgMYB4 plays a suppressive role in the regulation of anthocyanin synthesis during flower pigmentation.

Tulip bulbs are modified underground stems that originate from axillary meristems of mother bulb scales. Hormones, including jasmonic acids (JAs), play key roles in the regulation of tulip bulb development. Here, we compared variations in daughter bulb development through transcriptomic profiling analysis and characterized the functions of JA biosynthesis-related genes during daughter bulb enlargement. The results showed that tulip cultivars exhibited contrasting bulb size variations. Transcriptomic analyses revealed that genes involved in plant hormones and development, including the two lipoxygenase genes TgLOX4 and TgLOX5, showed significant changes in expression following tulip bulb growth. Ectopic overexpression of TgLOX4 and TgLOX5 in Arabidopsis enhanced endogenous JA content, improved plant growth, and increased lateral root numbers. Silencing of these two genes in tulip repressed the growth of daughter bulbs. Furthermore, exogenous JA treatment promoted tulip bulb growth, whereas the JA biosynthesis inhibitor sodium diethyldithiocarbamate (DIECA) inhibited this process. This study offers supporting evidence for the involvement of tulip TgLOX4 and TgLOX5 in the regulation of daughter bulb growth and development.