In China, Prunus mume is a famous flowering tree that has been cultivated for 3000 years. P. mume grows in tropical and subtropical regions, and most varieties lack cold resistance; thus, it is necessary to study the low-temperature response mechanism of P. mume to expand the scope of its cultivation. We used the integrated transcriptomic and metabolomic analysis of a cold-resistant variety of P. mume ‘Meiren’, to identify key genes and metabolites associated with low temperatures during flowering. The ‘Meiren’ cultivar responded in a timely manner to temperature by way of a low-temperature signal transduction pathway. After experiencing low temperatures, the petals fade and wilt, resulting in low ornamental value. At the same time, in the cold response pathway, the activities of related transcription factors up- or downregulate genes and metabolites related to low temperature-induced proteins, osmotic regulators, protective enzyme systems, and biosynthesis and metabolism of sugars and acids. Our findings promote research on the adaptation of P. mume to low temperatures during wintering and early flowering for domestication and breeding.

Osmanthus fragrans is an aromatic plant which is widely used in landscaping and garden greening in China. However, the process of flower opening is significantly affected by ambient temperature changes. Cell expansion in petals is the primary factor responsible for flower opening. Xyloglucan endoglycolase/hydrolase (XTH) is a cell-wall-loosening protein involved in cell expansion or cell-wall weakening. Through whole-genome analysis, 38 OfXTH genes were identified in O. fragrans which belong to the four main phylogenetic groups. The gene structure, chromosomal location, synteny relationship, and cis-acting elements prediction and expression patterns were analyzed on a genome-wide scale. The expression patterns showed that most OfXTHs were closely associated with the flower-opening period of O. fragrans. At the early flower-opening stage (S1 and S2), transcriptome and qRT-PCR analysis revealed the expression of OfXTH24, 27, 32, 35, and 36 significantly increased under low ambient temperature (19 °C). It is speculated that the five genes might be involved in the regulation of flower opening by responding to ambient temperature changes. Our results provide solid foundation for the functional analysis of OfXTH genes and help to explore the mechanism of flower opening responding to ambient temperature in O. fragrans.

In plants, the shikimate pathway is responsible for the production of aromatic amino acids L-tryptophan, L-phenylalanine, and L-tyrosine. L-Phenylalanine is the upstream substrate of flavonoid and anthocyanin synthesis. Shikimate kinase (SK) catalyzes the phosphorylation of the C3 hydroxyl group of shikimate to produce 3-phosphate shikimate (S3P), the fifth step of the shikimate pathway. However, whether SK participates in flavonoid and anthocyanin synthesis is unknown. This study characterized the single-copy PhSK gene in the petunia (Petunia hybrida) genome. PhSK was localized in chloroplasts. PhSK showed a high transcription level in corollas, especially in the coloring stage of flower buds. Suppression of PhSK changed flower color and shape, reduced the content of anthocyanins, and changed the flavonoid metabolome profile in petunia. Surprisingly, PhSK silencing caused a reduction in the shikimate, a substrate of PhSK. Further qPCR analysis showed that PhSK silencing resulted in a reduction in the mRNA level of PhDHQ/SDH, which encodes the protein catalyzing the third and fourth steps of the shikimate pathway, showing a feedback regulation mechanism of gene expression in the shikimate pathway.

Short-term ambient low temperature (ALT) stimulation is necessary for Osmanthus fragrans to facilitate continued flower opening after floral bud development reaches maturity. DNA methylation, a vital epigenetic modification, regulates various biological processes in response to temperature fluctuations. However, its role in temperature-driven flower opening remains elusive. In this study, we identified the pivotal timeframe during which O. fragrans promptly detected temperature cues. Using whole-genome bisulfite sequencing, we explored global DNA hypomethylation during this phase, with the most significant changes occurring in CHH sequence contexts. Auxin transport inhibitor (TIBA) application revealed that ALT-induced endogenous auxin accumulation promoted peduncle elongation. In our mRNA-seq analysis, we discovered that the differentially expressed genes (DEGs) with hypo-differentially methylated regions (hypo-DMRs) were mainly enriched in auxin and temperature response, RNA processing, and carbohydrate and lipid metabolism. Transcripts of three DNA demethylase genes (OfROS1a, OfDML3, OfDME) showed upregulation. Furthermore, all DNA methylase genes, except OfCMT2b, also displayed increased expression, specifically with two of them, OfCMT3a and OfCMT1, being associated with hypo-DMRs. Promoter assays showed that OfROS1a, with promoters containing low-temperature- and auxin-responsive elements, were activated by ALT and exogenous IAA at low concentrations but inhibited at high concentrations. Overexpression of OfROS1 reduced endogenous auxin levels but enhanced the expression of genes related to auxin response and spliceosome in petunia. Furthermore, OfROS1 promoted sucrose synthesis in petunia corollas. Our data characterized the rapid response of active DNA hypomethylation to ALT and suggested a possible epiregulation of temperature-dependent flower opening in O. fragrans. This study revealed the pivotal role of DNA hypomethylation in O. fragrans during the ALT-responsive phase before flower opening, involving dynamic DNA demethylation, auxin signaling modulation, and a potential feedback loop between hypomethylation and methylation.