Anthocyanins are a class of water-soluble flavonoid pigments that play crucial roles in plant physiology and human health. Their biosynthesis is regulated by the MYB-bHLH-WD40 transcriptional complex, in which MYB transcription factors serve as the key determinants. However, the function of repressor-type MYBs in tuber crops such as potato remains poorly understood. Here we show that the R2R3-MYB transcription factor StMYB3 acts as a repressor of anthocyanin biosynthesis in potato. Transient-expression assays revealed that co-expression of StMYB3 with StAN2 (an R2R3 MYB activator) markedly reduced anthocyanin accumulation in leaves, and tuber skins of StMYB3-over-expressing lines exhibited a significant decrease in anthocyanin content. Mechanistic analyses demonstrate that both StMYB3 and StAN2 interact with StAN1 (an anthocyanin related bHLH). StMYB3 not only suppresses the StAN2-mediated transcriptional activation of anthocyanin biosynthesis genes (StCHS, StF3H, StF3'5'H, and StGST) but also directly represses the promoter activity of bHLH transcription factor StAN1. Moreover, StMYB3 and StAN2 form a mutually reinforcing positive-feedback loop at the transcriptional level. These results uncovers a StMYB3-StAN1-StAN2 regulatory module whose dynamics, driven by competition and bidirectional feedback, precisely control anthocyanin accumulation. This finding provides a reference for breeding potatoes with high anthocyanin content.

Potato, the fourth largest food crop in the world, stores nutrients in underground tubers. However, light exposure induces tuber accumulation of chlorophyll and toxic steroidal glycoalkaloids (SGAs); this unwanted trait called tuber greening causes a decline in potato quality and renders parts of tuber inedible. Despite progress in enzymatic cascades governing SGAs biosynthesis, the regulatory scheme of SGAs and chlorophyll metabolism in light-exposed tubers persists as a critical knowledge gap. Here, we identify that the blue light receptor StCRY1 plays a predominant role in light-induced tuber greening and SGA elevation, functioning as a light-controlled transcriptional switch for genes involved in SGAs and chlorophyll biosynthesis. We show that the transcription factor StHY5 acts downstream of StCRY1 to coregulate both chlorophyll and SGA metabolism. However, StMYB4, a transcription factor regulated by StHY5, promotes SGA synthesis by directly binding to SGA biosynthetic genes without affecting chlorophyll homeostasis. Furthermore, StCRY1 employs a dual strategy by modulating StMYB4 expression and physically interacting with StMYB4 to regulate its transcriptional activity. Collectively, these findings uncover a modularly coordinated control of SGA accumulation and chlorophyll biosynthesis by bifurcation of the StCRY1-StHY5 axis, providing a promising strategy to concurrently curb light-induced tuber greening and glycoalkaloids accumulation.

The pigmentation patterns of potato tubers are complex and diverse, often exhibiting significant tissue specificity. This study was conducted to elucidate the molecular mechanisms underlying the differential pigmentation in different parts of potato tubers using two cultivars, ‘Huashu 12’ and 15EM36-26, which exhibit opposite pigmentation patterns between the bud eyes and the tuber periderm. Metabolomic analysis revealed that cyanidin, pelargonidin, and malvidin are the key anthocyanin components responsible for the observed pigmentation differences. A total of 118 common differentially expressed genes in the differentially pigmented tissues of both cultivars were identified in transcriptomic analysis, including key structural genes of the anthocyanin biosynthesis pathway (such as StPAL, StCHS, and StDFR). Weighted gene co-expression network analysis was further employed to screen modules significantly correlated with pigmentation phenotypes, and 28 candidate genes associated with anthocyanin biosynthesis were identified. Expression validation demonstrated that the expression of StbHLH14 was significantly higher in non-pigmented tissues compared to pigmented tissues. Functional analysis revealed that StbHLH14 can inhibit the activation of structural gene promoters (such as StCHS and StDFR) via the MYB transcription factor StAN2, thereby negatively regulating anthocyanin biosynthesis. This study unveils the metabolic and transcriptional basis of tissue-specific pigmentation in potato tubers and clarifies the negative regulatory role of StbHLH14.

Light is a key factor for inducing anthocyanin biosynthesis; however, its regulatory mode in potato anthocyanin biosynthesis remains unclear. Previous research identified a specific genotype that causes the tuber skin to gradually turn purple when exposed to light of different wavelengths. In the present study, we conducted metabolome and transcriptome analyses on tuber samples during anthocyanin accumulation. The metabolome data showed that the contents of naringenin chalcone, naringenin, dihydrokaempferol, and cyanidin gradually increased during anthocyanin accumulation. The transcriptome data showed that the expression levels of most structural genes increased gradually during anthocyanin accumulation, especially the StF3'H gene that promotes cyanidin formation. Moreover, the photo-responsive transcription factor StHY5 was specifically expressed at high levels before anthocyanin accumulation, occurring 2 h after light induction. Establishment of transgenic lines demonstrated that StHY5 overexpression could promote the accumulation of anthocyanin in potato tubers, along with a parallel increase in the transcription levels of StAN2, StMYBA1, StCHI, StF3H, StF3'H, and StDFR. Electrophoretic mobility shift and dual luciferase assays showed that StHY5 can enhance the promoter activity of the MYB transcription factors StAN2 and StMYBA1 as well as the structural genes StCHI and StF3H through binding to the G-box motif. StAN2 activated the expression of StF3'H (a newly identified purple gene locus in potato) and StDFR by binding to the MYB-binding site in the promoters, thereby promoting anthocyanin biosynthesis. This study provides a theoretical basis for revealing the molecular mechanism of light-regulated anthocyanin biosynthesis in potatoes.

During harvesting, storage, transportation, and processing, potato ( Solanum tuberosum L . ) tubers undergo greening after exposure to light, leading to the accumulation of toxic glycoside alkaloids, resulting in quality deterioration and economic losses. However, the underlying mechanisms are unclear. This study compared the transcriptome and proteome differences among four potato cultivars during the light-induced greening process, identifying 3,751 unique proteins (high confidence; ≥91.7%). The levels of enzymes involved in steroidal glycoalkaloid biosynthesis varied among the cultivars. In addition, coexpression network analysis of the transcriptomic data identified the transcription factor MYB113 (Soltu.DM.10G020780.1) as a potential positive regulator of steroidal glycoalkaloid biosynthesis. The dual-luciferase assay revealed that StMYB113 could bind to the promoters of steroidal glycoalkaloid biosynthesis-related genes and activate them. The transgenic lines overexpressing Solanum tuberosum L. Myb domain protein ( StMYB113 ) exhibited greater mRNA abundance of these genes and elevated levels of steroidal glycoalkaloids. This study provided a theoretical basis for exploring the impact of light on the synthesis of solanine in potatoes.

MYB transcription factors play a key role in anthocyanin biosynthesis. However, there are few reports on MYB transcription factors that can regulate anthocyanin biosynthesis in potato. In this study, the StMYB113 gene was isolated from potato and characterized. Transient activation experiments showed that the StMYB113 gene could promote anthocyanin biosynthesis in tobacco leaves by enhancing the expression of genes encoding anthocyanin biosynthesis-related enzymes. In addition, overexpression of the StMYB113 gene promoted anthocyanin accumulation in potato tuber skin. Furthermore, transient luciferase analysis showed that StMYB113 enhanced the promoter activities of StF3H and StDFR genes significantly. These results indicated that the StMYB113 gene could promote anthocyanin accumulation in potato tuber skin by directly activating the expression of StF3H and StDFR. The promoter activity of StMYB113 was significantly higher in light than in dark conditions, suggesting that the expression of StMYB113 is induced by light. These findings illustrate the role and regulation of the StMYB113 gene in potato anthocyanin biosynthesis and enrich the regulatory theory of potato anthocyanin biosynthesis.

Anthocyanin biosynthesis is affected by light, temperature, and other environmental factors. The regulation mode of light on anthocyanin synthesis in apple, pear, tomato and other species has been reported, while not clear in potato. In this study, potato RM-210 tubers whose peel will turn purple gradually after exposure to light were selected. Transcriptome analysis was performed on RM-210 tubers during anthocyanin accumulation. The expression of StMYBA1 gene continued to increase during the anthocyanin accumulation in RM-210 tubers. Moreover, co-expression cluster analysis of differentially expressed genes showed that the expression patterns of StMYBA1 gene were highly correlated with structural genes CHS and CHI . The promoter activity of StMYBA1 was significantly higher in light conditions, and StMYBA1 could activate the promoter activity of structural genes StCHS , StCHI , and StF3H . Further gene function analysis found that overexpression of StMYBA1 gene could promote anthocyanin accumulation and structural gene expression in potato leaves. These results demonstrated that StMYBA1 gene promoted potato anthocyanin biosynthesis by activating the expression of structural genes under light conditions. These findings provide a theoretical basis and genetic resources for the regulatory mechanism of potato anthocyanin synthesis.

C-repeat binding factors (CBFs) are well-known transcription factors (TFs) that regulate plant cold acclima-tion. RNA sequencing (RNA-seq) data from diverse plant species provide opportunities to identify other TFs involved in the cold response. However, this task is challenging because gene gain and loss has led to an intertwined community of co-orthologs and in-paralogs between and within species. Using orthogroup (closely related homologs) analysis, we identified 10,549 orthogroups in five representative eudicots. A phylotranscriptomic analysis of cold-treated seedlings from eudicots identified 35 high-confidence conserved cold-responsive transcription factor orthogroups (CoCoFos). These 35 CoCoFos included the well-known cold-responsive regulators CBFs, HSFC1, ZAT6/10, and CZF1 among others. We used Arabi-dopsis BBX29 for experimental validation. Expression and genetic analyses showed that cold-induction of BBX29 is CBF-and abscisic acid-independent, and BBX29 is a negative regulator of cold tolerance. Inte-grative RNA-seq and Cleavage Under Targets and Tagmentation followed by sequencing analyses revealed that BBX29 represses a set of cold-induced TFs (ZAT12, PRR9, RVE1, MYB96, etc.). Altogether, our analysis yielded a library of eudicot CoCoFos and demonstrated that BBX29 is a negative regulator of cold tolerance in Arabidopsis.

Angiosperms are one of the most diverse and abundant plant groups that are widely distributed on Earth, from tropical to temperate and polar zones. The wide distribution of angiosperms may be attributed to the evolution of sophisticated mechanisms of environmental adaptability, including cold tolerance. Since the development of high-throughput sequencing, transcriptome has been widely utilized to gain insights into the molecular mechanisms of plants in response to cold stress. However, previous studies generally focused on single or two species, and comparative transcriptome analyses for multispecies responding to cold stress were limited. In this study, we selected 11 representative angiosperm species, performed phylotranscriptome experiments at four time points before and after cold stress, and presented a profile of cold-induced transcriptome changes in angiosperms. Our multispecies cold-responsive RNA-seq datasets provide valuable references for exploring conserved and evolutionary mechanisms of angiosperms in adaptation to cold stress.

In the current potato germplasm resource preservation landscape,in vitro methods have gained widespread acceptance.Exploring the preservation conditions for extending the subculture cycle of test-tube seedlings can effectively reduce the risk and preservation cost.In this study,six different genotypes of potato materials were selected from the germplasm resources,and the effects of pre-rooting,temperature,sucrose concentration and subculture method on the preservation of potato test-tube seedlings were investigated using plant status,plant height and survival rate as indicators to explore the widely applicable conditions for long-term low temperature preservation of potato test-tube seedlings.Test-tube seedlings without pre-rooting treatment showed minimal growth on the 60th day of low temperature preservation and eventually died,suggesting that pre-rooting treatment is essential for the successful low temperature preservation of test-tube seedlings.The growth rate of test-tube seedlings at 4℃was slower compared to those at 7℃,and the plant height was obviously smaller on the 90th day.However,the survival rate of test-tube seedlings from four different materials at 7℃was 100%,while it was only 53.75%at 4℃,indicating that a temperature of 7℃is more suitable for preserving potato germplasm resources with various genotypes.Furthermore,the plant height of test-tube seedlings in a medium with 8%sucrose on the 90th day was obviously lower than those with 4%sucrose.Survival statistics revealed that all test-tube seedlings in the medium with 4%sucrose died after being stored at a temperature of 7℃for 270 days,whereas there was still a survival rate of approximately 61.25%for those in an 8%sucrose concentration medium.These findings suggest that an 8%sucrose concentration could prolong the low temperature preservation time for potato test-tube seedlings.In short,using a sucrose concentration of 8%in combination with an ambient temperature of 7℃could be effective for long-term low temperature preservation of most potato lines.Additionally,a subculture method involving test tube culture seeding and alternate box culture seedling was explored,which effectively improved the preservation of weak-growth germplasm resources and increased their survival rate from 22.50%(test tube culture seeding)to 95.00%(alternate subculture of test-tube culture seedlings-box culture seedlings)on the 90th day of low temperature preservation.This research provides valuable guidance for optimizing the conditions required for long-term preservation of potato germplasm resources.

Light-induced greening of tubers seriously affects its safety and economic benefits, but the mechanism of light-induced chlorophyll synthesis in potato tubers remains unclear. In this study, the related metabolites of potato tubers with different light durations were analyzed. The results were as follows: When the light duration prolonged, the chlorophyll content of the tubers gradually increased. Moreover, the chlorophyll content increased significantly at 36 hours, with which the tuber skin turned green obviously. Transcriptome sequencing and bioinformatics analysis were carried out on the samples taken at 0 hour, 6 hours, and 36hours, and 5646 differentially expressed genes（DEGs）were identified. According to the co-expression cluster analysis and quantitative RT-PCR verification results, 9 major structure genes of chlorophyll biosynthesis pathway（StGAS1, St CHLD, StCrd1,StHEMA, StGUN4, St PORA, St UROD, StCHLM, and StCHLG）and 6 transcription factors（St SBP, StLSD, St GATA, StWRKY,StMYB-like, and StMYB113）were significant induced. Predictions of the cis-acting elements of the promoter sequences of these 9structural genes indicated that they all contained multiple MYB binding sites. Promoter element analysis and transcriptional activation validation further revealed that StMYB113 had a light-responsive element and could activate the expression of StUROD in tobacco. Therefore, StMYB113 may be light-responsive and be able to regulate potato tuber greening in the light. This study provides a reference for the research on the regulatory mechanism of light-induced chlorophyll synthesis in potato tubers, which is important for reducing the loss caused by greening of potato tubers.

After anthocyanin synthesis, a variety of anthocyanin compounds are produced through further methylation, glycosylation, and acylation. However, the effect of the potato methylase gene on anthocyanin biosynthesis has not been reported. Red and purple mutation types appear in tubers of the potato cultivar 'Purple Viking' with chimeric skin phenotypes. In this study, transcriptome and anthocyanin metabolome analyses were performed on skin of Purple Viking tubers and associated mutants. According to the metabolome analysis, the transformation of delphinidin into malvidin-3-O-glucoside and petunidin 3-O-glucoside and that of cyanidin into rosinidin O-hexoside and peonidin-3-O-glucoside were hindered in red tubers. Expression of methyltransferase gene OMT30376 was significantly lower in red tubers than in purple ones, whereas the methylation level of OMT30376 was significantly higher in red tubers. In addition, red skin appeared in tubers from purple tuber plants treated with S-adenosylmethionine (SAM), indicating the difference between purple and red was caused by the methylation degree of the gene OMT30376. Thus, the results of the study suggest that the OMT30376 gene is involved in the transformation of anthocyanins in potato tubers. The results also provide an important reference to reveal the regulatory mechanisms of anthocyanin biosynthesis and transformation.

Long non-coding RNAs (lncRNAs) are emerging as versatile regulators in diverse biological processes. However, little is known about their cis- and trans-regulatory contributions in gene expression under salt stress. Using 27 RNA-seq data sets from Populus trichocarpa leaves, stems and roots, we identified 2988 high-confidence lncRNAs, including 1183 salt-induced differentially expressed lncRNAs. Among them, 301 lncRNAs have potential for positively affecting their neighboring genes, predominantly in a cis-regulatory manner rather than by co-transcription. Additionally, a co-expression network identified six striking salt-associated modules with a total of 5639 genes, including 426 lncRNAs, and in these lncRNA sequences, the DNA/RNA binding motifs are enriched. This suggests that lncRNAs might contribute to distant gene expression of the salt-associated modules in a trans-regulatory manner. Moreover, we found 30 lncRNAs that have potential to simultaneously cis- and trans-regulate salt-responsive homologous genes, and Ptlinc-NAC72, significantly induced under long-term salt stress, was selected for validating its regulation of the expression and functional roles of the homologs PtNAC72.A and PtNAC72.B (PtNAC72.A/B). The transient transformation of Ptlinc-NAC72 and a dual-luciferase assay of Ptlinc-NAC72 and PtNAC72.A/B promoters confirmed that Ptlinc-NAC72 can directly upregulate PtNAC72.A/B expression, and a presence/absence assay was further conducted to show that the regulation is probably mediated by Ptlinc-NAC72 recognizing the tandem elements (GAAAAA) in the PtNAC72.A/B 5' untranslated region (5'-UTR). Finally, the overexpression of Ptlinc-NAC72 produces a hypersensitive phenotype under salt stress. Altogether, our results shed light on the cis- and trans-regulation of gene expression by lncRNAs in Populus and provides an example of long-term salt-induced Ptlinc-NAC72 that could be used to mitigate growth costs by conferring plant resilience to salt stress.

Anthocyanins are biosynthesized on the cytoplasmic surface of the endoplasmic reticulum and then transported to the vacuole for storage. Glutathione S -transferases (GSTs) are involved in the transport of anthocyanins. However, the regulatory mechanism of GSTs in potato is unclear. In this study, the glutathione S -transferase gene StGST1 was isolated from potato and characterized. Expression analysis revealed that StGST1 transcript levels were higher in pigmented tubers than in white tubers. Overexpression of StGST1 promoted anthocyanin accumulation in the tuber. Transient luciferase analysis revealed that the promoter of StGST1 was activated by the transcription factors StAN2 and StAN11 . These results indicated that StGST1 promotes anthocyanin accumulation in potato tubers. These findings have important implications for improvement of anthocyanin accumulation in pigmented potato tubers.

The TEOSINTE BRANCHED1/CYCLOIDEA/PROLIFERATING CELL FACTOR (TCP) family proteins are plant-specific transcription factors that have been well-acknowledged for designing the architectures of plant branch, shoot, and inflorescence. However, evidence for their innovation and emerging role in abiotic stress has been lacking. In this study, we identified a total of 36 TCP genes in Populus trichocarpa, 50% more than that in Arabidopsis (i.e., 24). Comparative intra-genomes showed that such significant innovation was mainly due to the most recent whole genome duplication (rWGD) in Populus lineage around Cretaceous-Paleogene (K-Pg) boundary after the divergence from Arabidopsis. Transcriptome analysis showed that the expressions of PtrTCP genes varied among leaf, stem, and root, and they could also be elaborately regulated by abiotic stresses (e.g., cold and salt). Moreover, co-expression network identified a cold-associated regulatory module including PtrTCP31, PtrTCP10, and PtrTCP36. Of them, PtrTCP10 was rWGD-duplicated from PtrTCP31 and evolved a strong capability of cold induction, which might suggest a neofunctionalization of PtrTCP genes and contribute to the adaptation of Populus lineage during the Cenozoic global cooling. Evidentially, overexpression of PtrTCP10 into Arabidopsis increased freezing tolerance and salt susceptibility. Integrating co-expression network and cis-regulatory element analysis confirmed that PtrTCP10 can regulate the well-known cold- and salt-relevant genes (e.g., ZAT10, GolS2, and SOS1), proving that PtrTCP10 is an evolutionary innovation in P. trichocarpa response to environmental changes. Altogether, our results provide evidence of the rWGD in P. trichocarpa responsible for the innovation of PtrTCP genes and their emerging roles in environmental stresses.

Tuber shape is one of the most important traits for potato breeding. Since poor or irregular shape increases the difficulty of handling and processing, researching the inheritance of potato tuber shape for potato breeding is highly important. To efficiently identify QTL for tuber shape, a diploid potato population (PM7) was generated by self-pollinated M6 (S. chacoense). A QTL TScha6 for tuber shape was identified by the QTL-seq approach at 50.91-59.93 Mb on chromosome 6 in the potato DM reference genome. To confirm TScha6, four SSR and twenty CAPS markers around the QTL were developed and the TScha6 was narrowed down to an interval of ~ 1.85 Mb. The CAPS marker C6-58.27_665 linked to TScha6 was then used to screen 86 potato cultivars and advanced breeding lines. The tuber length/width (LW) ratio was significantly correlated with the presence/absence of C6-58.27_665, and the correlation coefficient was r = 0.55 (p < 0.01). These results showed that C6-58.27_665 could be applied in marker-assisted selection (MAS) for tuber shape breeding in the future. Our research sets the important stage for the future cloning of the tuber shape gene and utilities of the marker in the breeding program.

Previous studies on anthocyanin synthesis in potato mainly focus on the functions of anthocyanin biosynthetic structural genes and members of the MYB-bHLH-WD40 (MBW) complex. In this study, a RAV transcription factor StRAV1 was cloned. Expression analysis showed that the transcription of StRAV1 in yellow/white potato genotypes was significantly higher than in pigmented types. A transient activation assay showed that StRAV1 could inhibit anthocyanin formation in tobacco leaves by decreasing the expression of genes encoding related enzymes. Overexpression of StRAV1 in potato, led to significantly lower anthocyanin accumulation than in control tubers. The StCHS, StANS and St3’5’H promoters were significantly inhibited by StRAV1. These results suggest that the StRAV1 gene negatively regulates anthocyanin synthesis in potato tubers by directly inhibiting the transcription of StCHS, StANS and StF3’5’H. This is the first report on the regulation of anthocyanin synthesis in potato by a RAV gene, thus contributing to a more detailed understanding of this process.

The Bashang area of Hebei Province is a high-quality potato producing area in China. However, the drought and excessive exploitation of groundwater resources in this area seriously limit the promotion of local potato production. To clarify the response characteristics of different genotypes of potato to water stress, the drought stress phenotypes of ’Jingzhangshu 1’, ’Jingzhangshu 2’, ’Jingzhangshu 3’, ’Jizhangshu 8’ and ’Jizhangshu 12’ at the budding were evaluated against the sensitive varieties ’Shepody’ under the condition of artificial water control in rainproof shed.The average tuber weight and yield of all varieties decreased under the drought stress condition, and the drought resistance coefficients of ’Jingzhangshu 1’, ’Jingzhangshu 2’ and ’Jizhangshu 12’ were 0.68, 0.68 and 0.67 respectively,so the varieties mentioned above are strong resistant to drought. The drought resistance coefficients of ’Jingzhangshu 3’and ’Jizhangshu 8’ were 0.63 and 0.60, respectively, and the two varieties are resistant to drought. The yield of’Shepody’ was seriously reduced, and the drought resistance coefficient was 0.45, suggesting that it is a drought sensitive variety. It was found that the net photosynthetic rate, stomatal conductance and transpiration rate of’Jingzhangshu 1’ and ’Shepody’ were significantly higher than those of other varieties under drought stress for 15 days.After 30 days of drought stress, all photosynthetic indexes of ’Shepody’ decreased sharply, and photosynthesis stagnated. Other drought resistant varieties（including ’Jingzhangshu 1’） still maintained certain stomatal conductivity and net photosynthetic rate, indicating that ’Jingzhangshu 1’ showed drought resistance characteristics different from other varieties. Further observation of root development characteristics showed that the root shoot ratio of’Jingzhangshu 1’ and ’Jizhangshu 12’ was obviously increased, and the root shoot ratio of ’Jingzhangshu 1’ increased much greater. The number of lateral roots per plant, and length of adventitious roots and lateral roots of ’Jingzhangshu 1’and ’Jizhangshu 12’ were significantly higher than those of ’Shepody’, and the number of adventitious roots and lateral roots of ’Jingzhangshu 1’ was significantly higher than that of ’Jizhangshu 12’. It could be seen from the comprehensive indicators that the drought resistance characteristics of different genotypes of potatoes are not completely the same.Among them, the drought resistance of ’Jingzhangshu 1’ benefits from its developed root system, while the drought resistance of ’Jingzhangshu 2’, ’Jingzhangshu 3’, ’Jizhangshu 8’ and ’Jizhangshu 12’ mainly comes from their rapid stomatal regulation ability.

Although the role of WRKY transcription factors (TFs) in colour formation has been reported in several species, their function in potato (Solanum tuberosum L.) anthocyanin biosynthesis remains unclear. In this study, the potato WRKY gene StWRKY13 was isolated and characterised. Expression analysis revealed a significantly higher StWRKY13 expression in chromatic tubers than in yellow ones. Transient activation assays showed that StWRKY13 could enhance the role of StAN2 in promoting anthocyanin biosynthesis in tobacco (Nicotiana tabacum L.). Over-expressing the StWRKY13 gene promoted anthocyanin biosynthesis in potato tubers. Further investigations indicated that StWRKY13 could interact with the StCHS, StF3H, StDFR, and StANS gene promoters and significantly enhance their activities. Our findings showed that StWRKY13 could promote anthocyanin biosynthesis by activating StCHS, StF3H, StDFR, and StANS transcription in potato tubers, thereby supporting the theoretical basis for anthocyanins formation in coloured potato tubers.

As harbingers of bursting growth, flower buds and leaf buds generally show similar surface morphologies but different structural and functional changes. Dioecious plants further generate four types of Female/Male Flower/Leaf Buds (FFB, FLB, MFB, and MLB), showing a complex regulation. However, little is known about their underlying molecular mechanisms. Here, we exemplify the woody dioecious Salix linearistipularis to investigate their morphological characteristics and potential molecular mechanisms by combining cytological, physiological, phenological, and transcriptomic datasets. First, FFB and MFB have simultaneous development dynamics and so do FLB and MLB. Interestingly, FLB and MLB show very similar expression profiles preparing for photosynthesis and stress-tolerance, whereas FFB and MFB show great similarities but also striking sexual differences. Comparing flower buds and leaf buds after their revival from dormancy shows different cold- and vernalization-responsive genes (e.g. SliVRN1, SliAGL19, and SliAGL24), implying different programming processes for dormancy breaking between the buds. Moreover, except SliAP3, the expression of ABCDE model genes is consistent with their roles in the buds, suggesting a conserved mechanism of flower development between dioecious Salix and hermaphrodite Arabidopsis. Finally, considering sex-associated genes (e.g. SliCLE25, SliTPS21, and SliARR9) on Salix chromosomes and other reports, we hypothesize a dynamic model of sex determination on chromosomes 15 and 19 in the last ancestor of Salix and Populus but evolutionarily on 15 in Salix after their divergence. Together, our study provides new insights into the molecular mechanisms of dioecious four-type buds by showing the genes involved in their development, dormancy breaking, flowering, and sexual association.