DNA methylation, an important epigenetic modification, regulates the expression of genes and is therefore involved in the transitions between floral developmental stages in flowering plants. To explore whether DNA methylation plays different roles in the floral development of individual male and female dioecious plants, we injected 5-azacytidine (5-azaC), a DNA methylation inhibitor, into the trunks of female and male basket willow (Salix viminalis L.) trees before flower bud initiation. As expected, 5-azaC decreased the level of DNA methylation in the leaves of both male and female trees during floral development; however, it increased DNA methylation in the leaves of male trees at the flower transition stage. Furthermore, 5-azaC increased the number, length and diameter of flower buds in the female trees but decreased these parameters in the male trees. The 5-azaC treatment also decreased the contents of soluble sugars, starch and reducing sugars in the leaves of the female plants, while increasing them in the male plants at the flower transition stage; however, this situation was largely reversed at the flower development stage. In addition, 5-azaC treatment decreased the contents of auxin indoleacetic acid (IAA) in both male and female trees at the flower transition stage. These results indicate that hypomethylation in leaves at the flower transition stage promotes the initiation of flowering and subsequent floral growth in Salix viminalis, suggesting that DNA methylation plays a similar role in vegetative–reproductive transition and early floral development. Furthermore, methylation changes during the vegetative–reproductive transition and floral development were closely associated with the biosynthesis, metabolism and transportation of carbohydrates and IAA. These results provide insight into the epigenetic regulation of carbohydrate accumulation.

The objective of this study was to illustrate the photosynthetic characteristics of current twigs of Salix matsudana Koidz., and clarify the effect of stem photosynthesis on the new organ development in cutting seedlings. Excised twigs were taken as the experimental samples. The response of the stem photosynthesis rate to increasing light intensity and the effective photochemical efficiency of the cross section of the twig were determined. Then, twigs were used as cuttings and exposed to 0, 20, and 100 μmol m⁻² s⁻¹ light intensities, respectively, to achieve distinctive stem photosynthetic rates. After 14 days of treatment, stem water and non-structural carbohydrate (NSC) content, as well as the biomass and carbon isotopic composition, of new organs in the cutting seedlings under different light treatments were examined. The results showed that the gross photosynthetic rate significantly increased within 400 μmol m⁻² s⁻¹ of light intensity, and the maximum rate was approximately 1.27 μmol m⁻² s⁻¹. The effective photochemical efficiency of the PSⅡ of the cortex was significantly higher than the inner tissues in the cross section of the twig. When twig cuttings were exposed to different light intensities, stem water and starch content, as well as bud and root biomass, were significantly higher in the cutting seedling subjected to 100 μmol m⁻² s⁻¹ than the case treated in darkness; however, the bud δ¹³C trend was the opposite. Stem photosynthesis played a positive role in the maintenance of stem water and starch supply for the cutting seedlings, and ¹³C depleted assimilates produced by stem photosynthesis contributed to bud biomass, revealing that stem photosynthesis promotes organ development in cutting seedlings of Salix matsudana.

As a short-rotation woody plant, Salix viminalis has the potential for phytoremediation of cadmium (Cd), but it has poor tolerance to high Cd concentrations. Melatonin (MEL), a candidate bio-promoter, was considered to play an active role in plant responses to Cd. However, the molecular mechanism by which MEL regulates metabolic processes in plants to defend against Cd stress remain unclear. Transcriptomics and global untargeted metabolomic sequencing were used to investigate the rapid response of S. viminalis to high Cd concentrations during initial growth stage after foliar application of MEL. Four treatments were set up in a pot experiment involving foliar application of MEL on the first day, followed by irrigation with a Cd solution the next day. Significant variations in the relevant defence genes and metabolites in leaves exposed to Cd were observed between willows treated with and without MEL. Foliar application of MEL upregulated sulphur metabolism-related genes such as methionine and S-adenosylmethionine synthases in leaves exposed to Cd; glutamine content, which is the key point of nitrogen assimilation, also increased. Additionally, glycolysis and sucrose metabolic genes, including hexokinase, sucrose synthase, invertase, and the inositol phosphate metabolic gene myo-inositol-1-phosphate synthase were also upregulated in leaves. Moreover, MEL also upregulated genes related to the synthesis of flavonoids, anthocyanins, and proanthocyanins in the leaves. These results demonstrated that MEL improved amino acid and saccharide metabolism in the leaves of S. viminalis in response to Cd. It also improved the antioxidant capacity and Cd tolerance in S. viminalis leaves by enhancing synthetic capacity of polyphenol compounds. MEL may be involved in processes of photorespiration, ethylene metabolism, GABA shunt, nitric oxide metabolism, osmotic adjustment, and the synthesis of glutathione and ascorbate in S. viminalis under Cd stress. This series of metabolic changes in S. viminalis occurred within 24 h of the foliar application of MEL, which provided a sufficient substrate for subsequent defence reactions to cope with Cd stress. Our findings will help elucidate the molecular mechanism by which MEL regulates metabolic processes in plants in response to Cd challenges and guide the application of MEL to improve Cd phytoremediation efficiency.