Dormancy-associated MADS-box Research Articles

Impacts of climate change on the transcriptional dynamics and timing of bud dormancy release in Yoshino‐cherry tree

Societal Impact StatementThe iconic Yoshino cherry tree in Japan is experiencing shifts in its blossom timing due to climate warming. To develop a genetically informed predictive model for bud dormancy release, we examined seasonal gene expression in Yoshino cherry trees at three different locations. Our experiments, coupled with the analysis of DORMANCY‐ASSOCIATED MADS‐box (DAM) genes, highlighted DAM4 as the most reliable indicator for the rate of bud dormancy release. Our study demonstrated that seasonal gene expression profiles serve as a valuable indicator for forecasting the timing of dormancy release, benefiting Japanese traditions and providing insights into the biological impacts of climate change.Summary The Yoshino cherry tree Cerasus × yedoensis ‘Somei‐yoshino’ stands out as an iconic springtime symbol in Japan. For the Yoshino cherry trees to bloom in the spring, dormant buds must undergo a period of exposure to low temperatures, allowing them to break dormancy. Key genes related to dormancy release, known as DORMANCY‐ASSOCIATED MADS‐box (DAM), have been extensively studied. However, it remains unclear how these genes function in natural environments to regulate the timing of bud dormancy release. To develop a genetically informed predictive model for bud dormancy release, we explored seasonal changes in genome‐wide gene expression profiles in the Yoshino cherry trees at three distinct sites in Japan. Five distinct genome‐wide transcription profiles, subjectively named as modes—early summer, summer, autumn, winter, and spring—were identified, with the winter and spring modes observed when the daily mean temperature was below approximately 10°C. Our experiments of bud dormancy release, along with the assessment of expression profiles of DAM genes, have revealed that among the six DAM genes, DAM4 expression profile is the most indicative of the rate of bud dormancy break. Our estimates suggest that, on average, the tree needs to be exposed to temperatures below 10.1°C for 61.1 days to suppress DAM4 expression to the threshold required for bud dormancy release. Our projections for the timing of bud dormancy release indicated a delay of approximately 2.3 days per decade from 1990 to 2020. Our study demonstrated that gene expression serves as a valuable indicator for forecasting the timing of dormancy release.

Dormancy regulator Prunus mume DAM6 promotes ethylene-mediated leaf senescence and abscission.

Leaf senescence and abscission in autumn are critical phenological events in deciduous woody perennials. After leaf fall, dormant buds remain on deciduous woody perennials, which then enter a winter dormancy phase. Thus, leaf fall is widely believed to be linked to the onset of dormancy. In Rosaceae fruit trees, DORMANCY-ASSOCIATED MADS-box (DAM) transcription factors control bud dormancy. However, apart from their regulatory effects on bud dormancy, the biological functions of DAMs have not been thoroughly characterized. In this study, we revealed a novel DAM function influencing leaf senescence and abscission in autumn. In Prunus mume, PmDAM6 expression was gradually up-regulated in leaves during autumn toward leaf fall. Our comparative transcriptome analysis using two RNA-seq datasets for the leaves of transgenic plants overexpressing PmDAM6 and peach (Prunus persica) DAM6 (PpeDAM6) indicated Prunus DAM6 may up-regulate the expression of genes involved in ethylene biosynthesis and signaling as well as leaf abscission. Significant increases in 1-aminocyclopropane-1-carboxylate accumulation and ethylene emission in DEX-treated 35S:PmDAM6-GR leaves reflect the inductive effect of PmDAM6 on ethylene biosynthesis. Additionally, ethephon treatments promoted autumn leaf senescence and abscission in apple and P. mume, mirroring the changes due to PmDAM6 overexpression. Collectively, these findings suggest that PmDAM6 may induce ethylene emission from leaves, thereby promoting leaf senescence and abscission. This study clarified the effects of Prunus DAM6 on autumn leaf fall, which is associated with bud dormancy onset. Accordingly, in Rosaceae, DAMs may play multiple important roles affecting whole plant growth during the tree dormancy induction phase.

Functional evidence on the involvement of the MADS-box gene MdDAM4 in bud dormancy regulation in apple.

Over the course of the year, temperate trees experience extremes in temperature and day length. In order to protect themselves from frost damage in winter, they enter a dormant state with no visible growth where all leaves are shed and buds are dormant. Also the young floral tissues need to withstand harsh winter conditions, as temperature fruit trees like apple develop their flower buds in the previous year of fruit development. So far, the genetic control of induction and release of dormancy is not fully understood. However, the transcription factor family of DORMANCY-Associated MADS-box (DAM) genes plays a major role in the control of winter dormancy. One of these genes is MdDAM4. This gene is expressed in the early phase of bud dormancy, but little is known about its function. Six transgenic apple lines were produced to study the function of MdDAM4 in apple. For plant transformation, the binary plasmid vector p9oN-35s-MdDAM4 was used that contains the coding sequence of MdDAM4 driven by the 35S promoter. Transgenicity of the lines was proven by PCR and southern hybridization. Based on siRNA sequencing and phenotypic observations, it was concluded that line M2024 overexpresses MdDAM4 whereas the gene is silenced in all other lines. Phenotyping of the transgenic lines provided evidence that the overexpression of MdDAM4 leads to an earlier induction and a later release of dormancy. Silencing this gene had exactly the opposite effects and thereby led to an increased duration of the vegetation period. Expression experiments revealed genes that were either potentially repressed or activated by MdDAM4. Among the potentially suppressed genes were several homologs of the cytokinin oxidase 5 (CKX5), five LOX homologs, and several expansins, which may indicate a link between MdDAM4 and the control of leaf senescence. Among the potentially activated genes is MdDAM1, which is in line with observed expression patterns during winter dormancy. MdDAM2, which shows little expression during endodormancy also appears to be activated by MdDAM4. Overall, this study provides experimental evidence with transgenic apple trees for MdDAM4 being an important regulator of the onset of bud dormancy in apple.

Histone modifications affecting plant dormancy and dormancy release: common regulatory effects on hormone metabolism.

As sessile organisms, plants enter periods of dormancy in response to environmental stresses to ensure continued growth and reproduction in the future. During dormancy, plant growth is suppressed, adaptive/survival mechanisms are exerted, and stress tolerance increases over a prolonged period until the plants resume their development or reproduction under favorable conditions. In this review, we focus on seed dormancy and bud dormancy, which are critical for adaptation to fluctuating environmental conditions. We provide an overview of the physiological characteristics of both types of dormancy as well as the importance of the phytohormones abscisic acid and gibberellin for establishing and releasing dormancy, respectively. Additionally, recent epigenetic analyses have revealed that dormancy establishment and release are associated with the removal and deposition of histone modifications at the loci of key regulatory genes influencing phytohormone metabolism and signaling, including DELAY OF GERMINATION 1 and DORMANCY-ASSOCIATED MADS-box genes. We discuss our current understanding of the physiological and molecular mechanisms required to establish and release seed dormancy and bud dormancy, while also describing how environmental conditions control dormancy depth, with a focus on the effects of histone modifications.

Regulatory role of Prunus mume DAM6 on lipid body accumulation and phytohormone metabolism in the dormant vegetative meristem.

Bud dormancy is a crucial process in the annual growth cycle of woody perennials. In Rosaceae fruit tree species, DORMANCY-ASSOCIATED MADS-box (DAM) transcription factor genes regulating bud dormancy have been identified, but their molecular roles in meristematic tissues have not been thoroughly characterized. In this study, molecular and physiological analyses of transgenic apple plants overexpressing the Japanese apricot DAM6 gene (PmDAM6) and Japanese apricot cultivars and F1 individuals with contrasting dormancy characteristics revealed the metabolic pathways controlled by PmDAM6. Our transcriptome analysis and transmission electron microscopy examination demonstrated that PmDAM6 promotes the accumulation of lipid bodies and inhibits cell division in the dormant vegetative meristem by down-regulating the expression of lipid catabolism genes (GDSL ESTERASE/LIPASE and OIL BODY LIPASE) and CYCLIN genes, respectively. Our findings also indicate PmDAM6 promotes abscisic acid (ABA) accumulation and decreases cytokinin (CTK) accumulation in vegetative buds by up-regulating the expression of the ABA biosynthesis gene ARABIDOPSIS ALDEHYDE OXIDASE and the CTK catabolism gene CYTOKININ DEHYDROGENASE, while also down-regulating the expression of the CTK biosynthesis genes ISOPENTENYL TRANSFERASE (IPT) and CYP735A. Additionally, PmDAM6 modulates gibberellin (GA) metabolism by up-regulating GA2-OXIDASE expression and down-regulating GA3-OXIDASE expression. Furthermore, PmDAM6 may also indirectly promote lipid accumulation and restrict cell division by limiting the accumulation of CTK and GA in buds. In conclusion, using our valuable genetic platform, we clarified how PmDAM6 modifies diverse cellular processes, including lipid catabolism, phytohormone (ABA, CTK, and GA) biosynthesis and catabolism, and cell division, in the dormant vegetative meristem.

Transcription factors BZR2/MYC2 modulate brassinosteroid and jasmonic acid crosstalk during pear dormancy.

Bud dormancy is an important physiological process during winter. Its release requires a certain period of chilling. In pear (Pyrus pyrifolia), the abscisic acid (ABA)-induced expression of DORMANCY-ASSOCIATED MADS-box (DAM) genes represses bud break, whereas exogenous gibberellin (GA) promotes dormancy release. However, with the exception of ABA and GA, the regulatory effects of phytohormones on dormancy remain largely uncharacterized. In this study, we confirmed brassinosteroids (BRs) and jasmonic acid (JA) contribute to pear bud dormancy release. If chilling accumulation is insufficient, both 24-epibrassinolide (EBR) and methyl jasmonic acid (MeJA) can promote pear bud break, implying that they positively regulate dormancy release. BRASSINAZOLE RESISTANT 2 (BZR2), which is a BR-responsive transcription factor, inhibited PpyDAM3 expression and accelerated pear bud break. The transient overexpression of PpyBZR2 increased endogenous GA, JA, and JA-Ile levels. In addition, the direct interaction between PpyBZR2 and MYELOCYTOMATOSIS 2 (PpyMYC2) enhanced the PpyMYC2-mediated activation of Gibberellin 20-oxidase genes PpyGA20OX1L1 and PpyGA20OX2L2 transcription, thereby increasing GA3 contents and accelerating pear bud dormancy release. Interestingly, treatment with 5 μm MeJA increased the bud break rate, while also enhancing PpyMYC2-activated PpyGA20OX expression and increasing GA3,4 contents. The 100 μm MeJA treatment decreased the PpyMYC2-mediated activation of the PpyGA20OX1L1 and PpyGA20OX2L2 promoters and suppressed the inhibitory effect of PpyBZR2 on PpyDAM3 transcription, ultimately inhibiting pear bud break. In summary, our data provide insights into the crosstalk between the BR and JA signaling pathways that regulate the BZR2/MYC2-mediated pathway in the pear dormancy release process.

Single-Bud Expression Analysis of Bud Dormancy Factors in Peach.

Transcriptomic and gene expression analysis have greatly facilitated the identification and characterization of transcriptional regulatory factors and effectors involved in dormancy progression and other physiological processes orchestrated during bud development in peach and other temperate fruit species. Gene expression measurements are most usually based on average values from several or many individual buds. We have performed single-bud gene analysis in flower buds of peach across dormancy release using amplicons from the master regulatory DORMANCY-ASSOCIATED MADS-BOX (DAM) factors, several jasmonic acid biosynthetic genes, other genes related to flowering development, cell growth resumption, and abiotic stress tolerance. This analysis provides a close view on gene-specific, single-bud variability throughout the developmental shift from dormant to dormancy-released stages, contributing to the characterization of putative co-expression modules and other regulatory aspects in this particular tissue.

Peculiarity of transcriptional and H3K27me3 dynamics during peach bud dormancy

Bud dormancy facilitates the survival of meristems under harsh environmental conditions. To elucidate how molecular responses to chilling accumulation controlling dormancy in peach buds, chromatin immunoprecipitation sequencing to identify the H3K27me3 modifications and RNA sequencing of two peach cultivars with pronounced differences in chilling requirement were carried out, the results showed that genes associated with abscisic acid and gibberellic acid signal pathways play key roles in dormancy regulation. The results demonstrated that peach flower bud differentiation occurred continuously in both cultivars during chilling accumulation, which was correlated with the transcript abundance of key genes involved in phytohormone metabolism and flower bud development under adverse conditions. The more increased strength in high chilling-requirement cultivar along with the chilling accumulation at the genome-wide level. The function of the dormancy-associated MADS-box gene PpDAM6 was identified, which is involved in leaf bud break in peach and flower development in transgenic Nicotiana tabacum (NC89). In addition, PpDAM6 was positively regulated by PpCBF, and the genes of putative dormancy-related and associated with metabolic pathways were proposed. Taken together, these results constituted a theoretical basis for elucidating the regulation of peach bud dormancy transition.

MADS-box protein PpDAM6 regulates chilling requirement-mediated dormancy and bud break in peach.

Bud dormancy is crucial for winter survival and is characterized by the inability of the bud meristem to respond to growth-promotive signals before the chilling requirement (CR) is met. However, our understanding of the genetic mechanism regulating CR and bud dormancy remains limited. This study identified PpDAM6 (DORMANCY-ASSOCIATED MADS-box) as a key gene for CR using a GWAS (Genome-Wide Association Study) analysis based on structural variations (SVs) in 345 peach (Prunus persica (L.) Batsch) accessions. The function of PpDAM6 in CR regulation was demonstrated by transiently silencing the gene in peach buds and stably overexpressing the gene in transgenic apple (Malus × domestica) plants. The results showed an evolutionarily conserved function of PpDAM6 in regulating bud dormancy release, followed by vegetative growth and flowering, in peach and apple. The 30-bp deletion in the PpDAM6 promoter was substantially associated with reducing PpDAM6 expression in low-CR accessions. A PCR marker based on the 30-bp indel was developed to distinguish peach plants with non-low and low CR. Modification of the H3K27me3 marker at the PpDAM6 locus showed no apparent change across the dormancy process in low- and non-low-chilling requirement cultivars. Additionally, H3K27me3 modification occurred earlier in low-CR cultivars on a genome-wide scale. PpDAM6 could mediate cell-cell communication by inducing the expression of the downstream genes PpNCED1 (9-cis-epoxycarotenoid dioxygenase 1), encoding a key enzyme for ABA biosynthesis, and CALS (CALLOSE SYNTHASE), encoding callose synthase. We shed light on a gene regulatory network formed by PpDAM6-containing complexes that mediate CR underlying dormancy and budbreak in peach. A better understanding of the genetic basis for natural variations of CR can help breeders develop cultivars with different CR for growing in different geographical regions.

The study of the characteristics of the secondary flowering of Cerasus subhirtella 'Autumnalis'.

The short flowering period of ornamental cherry trees is the main factor limiting their use in gardens. Determining the secondary flowering characteristics of ornamental cherry trees is required to prolong their flowering period. In this experiment, Cerasus subhirtella 'Autumnalis' was used as the experimental material. The phenological differences in their annual growth cycle were observed using the BBCH coding system. The cooling requirements of the flower buds were evaluated by the chilling hours model (temperature between 0 and 7.2 °C) and the Utah model. The expression of the core gene involved in bud dormancy regulation DAM (dormancy-associated MADS-box) from the completion of flower bud differentiation in one year until the following year was measured by performing real-time fluorescence-based quantitative PCR. The results showed that the flowering duration of C. subhirtella 'Autumnalis' from November to December was longer than that of C. yedoensis 'Somei Yoshino', which was from March to April. The progress from seed bud-break to flower bud opening took about 10 days for C. subhirtella 'Autumnalis', while the same stage for C. yedoensis 'Somei Yoshino' took around 20 days. Additionally, the flower buds of C. subhirtella 'Autumnalis' needed only the chilling temperature unit of 54.08 to satisfy the chilling requirement, while C. yedoensis 'Somei Yoshino' required a chilling temperature unit of 596.75. After the completion of flower bud differentiation, during low-temperature storage, the expression of DAM4 and DAM5 genes first increased and then decreased, whereas, the expression of the DAM6 gene continued to decrease, and the expression of DAM4, DAM5, and DAM6 in C. yedoensis 'Somei Yoshino' increased rapidly and was maintained at a high level. This showed that the upregulation of the expression of the DAM4, DAM5, and DAM6 genes can inhibit the flower bud germination of Cherry Blossom. The relative expression of the DAM gene of C. subhirtella 'Autumnalis' was significantly lower than that of the DAM gene of C. yedoensis 'Somei Yoshino' from the end of October to the beginning of December, leading to its secondary flowering in autumn. These results might elucidate why the flower buds of C. subhirtella 'Autumnalis' can break their internal dormancy and bloom in the autumn and then again in the following year. Our findings might provide a reference for conducting further studies on the mechanisms of secondary flowering and bud dormancy in cherries.

The collaborative mode by PmSVPs and PmDAMs reveals neofunctionalization in the switch of the flower bud development and dormancy for Prunus mume.

Prunus mume (Rosaceae, Prunoideae) serves as an excellent ornamental woody plant with a large-temperature-range cultivation scope. Its flower buds require a certain low temperature to achieve flowering circulation. Thus, it is important to delve into the processes of flower bud differentiation and dormancy, which affected its continuous flowering. These processes are generally considered as regulation by the MADS-box homologs, SHORT VEGETATIVE PHASE (SVP), and DORMANCY-ASSOCIATED MADS-BOX (DAM). However, a precise model on their interdependence and specific function, when acting as a complex in the flower development of P. mume, is needed. Therefore, this study highlighted the integral roles of PmDAMs and PmSVPs in flower organ development and dormancy cycle. The segregation of PmDAMs and PmSVPs in a different cluster suggested distinct functions and neofunctionalization. The expression pattern and yeast two-hybrid assays jointly revealed that eight genes were involved in the floral organ development stages, with PmDAM1 and PmDAM5 specifically related to prolificated flower formation. PmSVP1-2 mingled in the protein complex in bud dormancy stages with PmDAMs. Finally, we proposed the hypothesis that PmSVP1 and PmSVP2 could combine with PmDAM1 to have an effect on flower organogenesis and interact with PmDAM5 and PmDAM6 to regulate flower bud dormancy. These findings could help expand the current molecular mechanism based on MADS-box genes during flower bud development and dormancy.

Epigenetics for crop adaptation to climate change

Epigenetics for crop adaptation to climate change

Evolutionary origin and functional specialization of Dormancy-Associated MADS box (DAM) proteins in perennial crops

BackgroundBud dormancy is a phenological adaptation of temperate perennials that ensures survival under winter temperature conditions by ceasing growth and increasing cold hardiness. SHORT VEGETATIVE PHASE (SVP)-like factors, and particularly a subset of them named DORMANCY-ASSOCIATED MADS-BOX (DAM), are master regulators of bud dormancy in perennials, prominently Rosaceae crops widely adapted to varying environmental conditions.ResultsSVP-like proteins from recently sequenced Rosaceae genomes were identified and characterized using sequence, phylogenetic and synteny analysis tools. SVP-like proteins clustered in three clades (SVP1–3), with known DAM proteins located within SVP2 clade, which also included Arabidopsis AGAMOUS-LIKE 24 (AthAGL24). A more detailed study on these protein sequences led to the identification of a 15-amino acid long motif specific to DAM proteins, which affected protein heteromerization properties by yeast two-hybrid system in peach PpeDAM6, and the unexpected finding of predicted DAM-like genes in loquat, an evergreen species lacking winter dormancy. DAM gene expression in loquat trees was studied by quantitative PCR, associating with inflorescence development and growth in varieties with contrasting flowering behaviour.ConclusionsPhylogenetic, synteny analyses and heterologous overexpression in the model plant Arabidopsis thaliana supported three major conclusions: 1) DAM proteins might have emerged from the SVP2 clade in the Amygdaloideae subfamily of Rosaceae; 2) a short DAM-specific motif affects protein heteromerization, with a likely effect on DAM transcriptional targets and other functional features, providing a sequence signature for the DAM group of dormancy factors; 3) in agreement with other recent studies, DAM associates with inflorescence development and growth, independently of the dormancy habit.

H3K4me3 plays a key role in establishing permissive chromatin states during bud dormancy and bud break in apple.

Bud dormancy helps woody perennials survive winter and activate robust plant development in the spring. For apple (Malus × domestica), short-term chilling induces bud dormancy in autumn, then prolonged chilling leads to dormancy release and a shift to a quiescent state in winter, with subsequent warm periods promoting bud break in spring. Epigenetic regulation contributes to seasonal responses such as vernalization. However, how histone modifications integrate seasonal cues and internal signals during bud dormancy in woody perennials remains largely unknown. Here, we show that H3K4me3 plays a key role in establishing permissive chromatin states during bud dormancy and bud break in apple. The global changes in gene expression strongly correlated with changes in H3K4me3, but not H3K27me3. High expression of DORMANCY-ASSOCIATED MADS-box (DAM) genes, key regulators of dormancy, in autumn was associated with high H3K4me3 levels. In addition, known DAM/SHORT VEGETATIVE PHASE (SVP) target genes significantly overlapped with H3K4me3-modified genes as bud dormancy progressed. These data suggest that H3K4me3 contributes to the central dormancy circuit, consisting of DAM/SVP and abscisic acid (ABA), in autumn. In winter, the lower expression and H3K4me3 levels at DAMs and gibberellin metabolism genes control chilling-induced release of dormancy. Warming conditions in spring facilitate the expression of genes related to phytohormones, the cell cycle, and cell wall modification by increasing H3K4me3 toward bud break. Our study also revealed that activation of auxin and repression of ABA sensitivity in spring are conditioned at least partly through temperature-mediated epigenetic regulation in winter.

Populus SVL Acts in Leaves to Modulate the Timing of Growth Cessation and Bud Set.

SHORT VEGETATIVE PHASE (SVP) is an important regulator of FLOWERING LOCUS T (FT) in the thermosensory pathway of Arabidopsis. It is a negative regulator of flowering and represses FT transcription. In poplar trees, FT2 is central for the photoperiodic control of growth cessation, which also requires the decrease of bioactive gibberellins (GAs). In angiosperm trees, genes similar to SVP, sometimes named DORMANCY-ASSOCIATED MADS-BOX genes, control temperature-mediated bud dormancy. Here we show that SVL, an SVP ortholog in aspen trees, besides its role in controlling dormancy through its expression in buds, is also contributing to the regulation of short day induced growth cessation and bud set through its expression in leaves. SVL is upregulated during short days in leaves and binds to the FT2 promoter to repress its transcription. It furthermore decreases the amount of active GAs, whose downregulation is essential for growth cessation, by repressing the transcription of GA20 oxidase. Finally, the SVL protein is more stable in colder temperatures, thus integrating the temperature signal into the response. We conclude that the molecular function of SVL in the photoperiodic pathway has been conserved between Arabidopsis and poplar trees, albeit the physiological process it controls has changed. SVL is thus both involved in regulating the photoperiod response in leaves, modulating the timing of growth cessation and bud set, and in the subsequent temperature regulation of dormancy in the buds.

How Is Global Warming Affecting Fruit Tree Blooming? "Flowering (Dormancy) Disorder" in Japanese Pear (Pyrus pyrifolia) as a Case Study.

Recent climate change has resulted in warmer temperatures. Warmer temperatures from autumn to spring has negatively affected dormancy progression, cold (de)acclimation, and cold tolerance in various temperate fruit trees. In Japan, a physiological disorder known as flowering disorder, which is an erratic flowering and bud break disorder, has recently emerged as a serious problem in the production of the pome fruit tree, Japanese (Asian) pear (Pyrus pyrifolia Nakai). Due to global warming, the annual temperature in Japan has risen markedly since the 1990s. Surveys of flowering disorder in field-grown and greenhouse-grown Japanese pear trees over several years have indicated that flowering disorder occurs in warmer years and cultivation conditions, and the risk of flowering disorder occurrence is higher at lower latitudes than at higher latitudes. Susceptibility to flowering disorder is linked to changes in the transcript levels of putative dormancy/flowering regulators such as DORMANCY-ASSOCIATED MADS-box (DAM) and FLOWERING LOCUS T (FT). On the basis of published studies, we conclude that autumn–winter warm temperatures cause flowering disorder through affecting cold acclimation, dormancy progression, and floral bud maturation. Additionally, warm conditions also decrease carbohydrate accumulation in shoots, leading to reduced tree vigor. We propose that all these physiological and metabolic changes due to the lack of chilling during the dormancy phase interact to cause flowering disorder in the spring. We also propose that the process of chilling exposure rather than the total amount of chilling may be important for the precise control of dormancy progression and robust blooming, which in turn suggests the necessity of re-evaluation of the characteristics of cultivar-dependent chilling requirement trait. A full understanding of the molecular and metabolic regulatory mechanisms of both dormancy completion (floral bud maturation) and dormancy break (release from the repression of bud break) will help to clarify the physiological basis of dormancy-related physiological disorder and also provide useful strategies to mitigate or overcome it under global warming.

The genome of low-chill Chinese plum "Sanyueli" (Prunus salicina Lindl.) provides insights into the regulation of the chilling requirement of flower buds.

Chinese plum (Prunus salicina Lindl.) is a stone fruit that belongs to the Prunus genus and plays an important role in the global production of plum. In this study, we report the genome sequence of the Chinese plum "Sanyueli", which is known to have a low-chill requirement for flower bud break. The assembled genome size was 282.38Mb, with a contig N50 of 1.37Mb. Over 99% of the assembly was anchored to eight pseudochromosomes, with a scaffold N50 of 34.46Mb. A total of 29,708 protein-coding genes were predicted from the genome and 46.85% (132.32Mb) of the genome was annotated as repetitive sequence. Bud dormancy is influenced by chilling requirement in plum and partly controlled by DORMANCY ASSOCIATED MADS-box (DAM) genes. Six tandemly arrayed PsDAM genes were identified in the assembled genome. Sequence analysis of PsDAM6 in "Sanyueli" revealed the presence of large insertions in the intron and exon regions. Transcriptome analysis indicated that the expression of PsDAM6 in the dormant flower buds of "Sanyueli" was significantly lower than that in the dormant flower buds of the high chill requiring "Furongli" plum. In addition, PsDAM6 expression was repressed by chilling treatment. The genome sequence of "Sanyueli" plum provides a valuable resource for elucidating the molecular mechanisms responsible for the regulation of chilling requirements, and it is also useful for the identification of the genes involved in the control of other important agronomic traits and molecular breeding in plum.

Regulatory circuits involving bud dormancy factor PpeDAM6

DORMANCY-ASSOCIATED MADS-BOX (DAM) genes have recently emerged as key potential regulators of the dormancy cycle and climate adaptation in perennial species. Particularly, PpeDAM6 has been proposed to act as a major repressor of bud dormancy release and bud break in peach (Prunus persica). PpeDAM6 expression is downregulated concomitantly with the perception of a given genotype-dependent accumulation of winter chilling time, and the coincident enrichment in H3K27me3 chromatin modification at a specific genomic region. We have identified three peach BASIC PENTACYSTEINE PROTEINs (PpeBPCs) interacting with two GA-repeat motifs present in this H3K27me3-enriched region. Moreover, PpeBPC1 represses PpeDAM6 promoter activity by transient expression experiments. On the other hand, the heterologous overexpression of PpeDAM6 in European plum (Prunus domestica) alters plant vegetative growth, resulting in dwarf plants tending toward shoot meristem collapse. These alterations in vegetative growth of transgenic lines associate with impaired hormone homeostasis due to the modulation of genes involved in jasmonic acid, cytokinin, abscisic acid, and gibberellin pathways, and the downregulation of shoot meristem factors, specifically in transgenic leaf and apical tissues. The expression of many of these genes is also modified in flower buds of peach concomitantly with PpeDAM6 downregulation, which suggests a role of hormone homeostasis mechanisms in PpeDAM6-dependent maintenance of floral bud dormancy and growth repression.

Unraveling the role of MADS transcription factor complexes in apple tree dormancy.

Summary A group of MADS transcription factors (TFs) are believed to control temperature‐mediated bud dormancy. These TFs, called DORMANCY‐ASSOCIATED MADS‐BOX (DAM), are encoded by genes similar to SHORT VEGETATIVE PHASE (SVP) from Arabidopsis. MADS proteins form transcriptional complexes whose combinatory composition defines their molecular function. However, how MADS multimeric complexes control the dormancy cycle in trees is unclear.Apple MdDAM and other dormancy‐related MADS proteins form complexes with MdSVPa, which is essential for the ability of transcriptional complexes to bind to DNA. Sequential DNA‐affinity purification sequencing (seq‐DAP‐seq) was performed to identify the genome‐wide binding sites of apple MADS TF complexes. Target genes associated with the binding sites were identified by combining seq‐DAP‐seq data with transcriptomics datasets obtained using a glucocorticoid receptor fusion system, and RNA‐seq data related to apple dormancy.We describe a gene regulatory network (GRN) formed by MdSVPa‐containing complexes, which regulate the dormancy cycle in response to environmental cues and hormonal signaling pathways. Additionally, novel molecular evidence regarding the evolutionary functional segregation between DAM and SVP proteins in the Rosaceae is presented.MdSVPa sequentially forms complexes with the MADS TFs that predominate at each dormancy phase, altering its DNA‐binding specificity and, therefore, the transcriptional regulation of its target genes.

High-quality genome assembly of 'Cuiguan' pear (Pyrus pyrifolia) as a reference genome for identifying regulatory genes and epigenetic modifications responsible for bud dormancy

Dormancy-associated MADS-box (DAM) genes serve as crucial regulators of the endodormancy cycle in rosaceous plants. Although pear DAM genes have been identified previously, the lack of a high-quality reference genome and techniques to study gene function have prevented accurate genome-wide analysis and functional verification of such genes. Additionally, the contribution of other genes to the regulation of endodormancy release remains poorly understood. In this study, a high-quality genome assembly for 'Cuiguan' pear (Pyrus pyrifolia), which is a leading cultivar with a low chilling requirement cultivated in China, was constructed using PacBio and Hi-C technologies. Using this genome sequence, we revealed that pear DAM genes were tandemly clustered on Chr8 and Chr15 and were differentially expressed in the buds between 'Cuiguan' and the high-chilling-requirement cultivar 'Suli' during the dormancy cycle. Using a virus-induced gene silencing system, we determined the repressive effects of DAM genes on bud break. Several novel genes potentially involved in the regulation of endodormancy release were identified by RNA sequencing and H3K4me3 chromatin immunoprecipitation sequencing analyses of 'Suli' buds during artificial chilling using the new reference genome. Our findings enrich the knowledge of the regulatory mechanism underlying endodormancy release and chilling requirements and provide a foundation for the practical regulation of dormancy release in fruit trees as an adaptation to climate change.