Nucleoskeleton proteins modulate nuclear structure and play important roles in diverse cellular processes. Yet, their functions in immune responses are poorly understood. Here, we report that a nucleoskeleton protein, po...Nucleoskeleton proteins modulate nuclear structure and play important roles in diverse cellular processes. Yet, their functions in immune responses are poorly understood. Here, we report that a nucleoskeleton protein, potato lamin-like protein StKAKU4, is targeted and stabilized by the oomycete RxLR effector Pi22798 to suppress immunity. StKAKU4 predominantly localizes to the nuclear envelope (NE) and is frequently trafficked between the NE and endosomal organelles. Enhanced expression of KAKU4 in Nicotiana benthamiana significantly increases Phytophthora infestans infection, whereas silencing of KAKU4 in potato and N. benthamiana enhances resistance, suggesting that it acts as a susceptibility (S) factor. GFP-StKAKU4 accumulates at the extrahaustorial membrane (EHM) during P. infestans infection. Moreover, Pi22798 interacts with StKAKU4 at the NE and accelerates endomembrane trafficking of StKAKU4 to the cytoplasm. An endomembrane enrichment assay indicates that StKAKU4 is involved in intracellular transport. In the immunopurified GFP-StKAKU4-associated endomembrane proteome, both host immunity-associated proteins and several P. infestans RxLR effectors are enriched, whereas apoplastic effectors such as PiSCR74, which do not enter host cells, are not. Our findings reveal a mechanism by which the potato NE protein StKAKU4 is involved in vesicle trafficking and is exploited by P. infestans to facilitate infection and colonization, offering valuable insights into S factors and potential strategies for improving disease resistance.
Floral organ development is a crucial process in plant life, and its precise regulation is essential for crop yield. In this study, we demonstrate that BnaA02.TOP1α and BnaC02.TOP1α, type IB DNA topoisomerases, play a pi...Floral organ development is a crucial process in plant life, and its precise regulation is essential for crop yield. In this study, we demonstrate that BnaA02.TOP1α and BnaC02.TOP1α, type IB DNA topoisomerases, play a pivotal role in regulating floral organ development in Brassica napus. The double mutant of these two genes, s5, exhibited an increased probability of developing malformed floral organ structures, along with a reduced number of second and third whorl floral organs and an increased number of first and fourth whorl floral organs. These phenotypic defects were primarily attributed to the downregulation of the floral meristem identity genes BnaC03.LFY, BnaA08.UFO, and BnaC08.UFO, and to the altered expression of several B- and E-class genes in the s5 mutant. BnaA02.TOP1α physically bound to the genomic regions of BnaC03.LFY, BnaA08.UFO, and BnaC08.UFO, promoting the recruitment of the RNA polymerase II complex to their transcription start sites. These changes were associated with alterations in histone modifications and the accumulation of DNA:RNA hybrids. Our findings demonstrate that BnaA02.TOP1α regulates DNA topology, thereby facilitating the recruitment of RNA polymerase II at BnaC03.LFY, BnaA08.UFO, and BnaC08.UFO. In conjunction with histone modifications and R-loops, this recruitment promotes the expression of these genes, thereby ensuring normal floral organ development. These findings contribute to our understanding of BnaTOP1α function and provide valuable insights into the regulatory network controlling floral organ development in B. napus.
Wang S, Zhao X, Wang X
… +16 more, Wang M, Wang C, Liu M, Zhu Y, Liu T, Wang Y, Chen X, Xu Y, Cheng F, Che X, Liu X, Lv B, Chai S, Xu K, Zhang Z, Zhang H
Cucurbitacin B (CuB) is the primary compound responsible for bitterness in melon (Cucumis melo L.) and negatively affects fruit quality and consumer acceptance. Although CuB biosynthesis is regulated by multiple transcri...Cucurbitacin B (CuB) is the primary compound responsible for bitterness in melon (Cucumis melo L.) and negatively affects fruit quality and consumer acceptance. Although CuB biosynthesis is regulated by multiple transcription factors, the regulatory mechanisms underlying CuB biosynthesis across different tissues remain unclear. Here, we generated 2 independent knockout lines (Cmbt-18 and Cmbt-19) of the basic helix-loop-helix (bHLH) transcription factor gene CmBt (Bitter fruit) using CRISPR-Cas9. Loss of CmBt reduces CuB levels in fruits and roots and produces non-bitter leaves, indicating that CmBt predominantly regulates CuB accumulation in leaves. By contrast, its paralog CmBr (Bitter root) predominantly controls CuB accumulation in fruits and roots. These results redefine CmBt as a leaf-predominant regulator and CmBr as a fruit/root-predominant regulator of CuB biosynthesis, revising previous findings. Furthermore, we demonstrate that CmBt physically interacts with CmBr. The CmbtCmbr double mutants exhibit a more severe reduction in CuB content in fruits, leaves, and roots compared with either single mutant, confirming a synergistic interaction between CmBt and CmBr. Reverse transcription quantitative PCR and tobacco dual-luciferase assays further show that CmBt and CmBr synergistically activate the CuB biosynthetic gene CmBi. Additionally, the CmbtCmbr double mutant completely prevents CPPU-induced fruit bitterness. Our findings reveal that CmBt and CmBr exert tissue-specific, partially redundant, and synergistic roles in activating CmBi and promoting CuB biosynthesis. These results identify the CmBt-CmBr locus as a promising target for breeding non-bitter melon cultivars.
Advances in quantitative genetics have enabled researchers to identify genomic regions associated with changes in phenotype. However, these regions can contain hundreds to thousands of genes, and progressing from genomic...Advances in quantitative genetics have enabled researchers to identify genomic regions associated with changes in phenotype. However, these regions can contain hundreds to thousands of genes, and progressing from genomic regions to causative genes is still challenging. In genome-wide association studies (GWAS) measuring elemental accumulation (ionomic) traits, only 5% of loci contain orthologs of genes known to control the ionome - indicating that many causal genes are still unknown. To identify candidates for the remaining 95% of loci, we developed a method that uses GWAS studies across multiple species to identify conserved causative genes. By Filtering the Results of Multi-species, Analogous, GWAS Experiments (FiReMAGE), we processed the GWAS of 19 ionomic traits in Arabidopsis (Arabidopsis thaliana), soybean (Glycine max), rice (Oryza sativa), maize (Zea mays), and sorghum, and identified alleles affecting trait variation at genes conserved across these five species. Permutation testing demonstrated that homologous genes were present in loci affecting element accumulation more often than expected. The repeated recovery of homologs at GWAS loci highlights the conservation of ionomic genetic regulators in flowering plants. FiReMAGE identified more candidate genes encoding proteins with known roles in regulating the ionome than expected by chance, validating the approach. These alleles are directly available for improving plant nutrition, food nutritional value, and food safety. FiReMAGE also identified conserved genes with no previously identified function. This provides a path to discover mechanisms of trait variation and conserved genes of previously unknown function via GWAS. The scripts to run FiReMAGE and adapt it to any trait are available on GitHub.
Grain size is a crucial agronomic trait that influences the grain weight and yield of rice (Oryza sativa L.). Here, we identified and characterized OsPAR1, an HLH-type miP (microProtein), as a positive regulator of grain...Grain size is a crucial agronomic trait that influences the grain weight and yield of rice (Oryza sativa L.). Here, we identified and characterized OsPAR1, an HLH-type miP (microProtein), as a positive regulator of grain size and grain weight. Loss-of-function Ospar1 mutants exhibited reduced cell size of the rice spikelet hulls, resulting in significantly smaller grains and lower grain weight. Conversely, OsPAR1-overexpressing lines displayed enlarged hull cells, leading to considerably larger grains and increased grain weight. Biochemical analyses showed that OsPAR1 interacts with the bHLH transcription factor OsBCL1 and functions as a transcriptional co-activator. OsPAR1 promotes OsBCL1 binding to the promoter of OsEXPA5, a key gene involved in cell expansion. Meanwhile, Osbcl1 and Osexpa5 mutants showed smaller grains with decreased grain length, grain width, grain thickness, and 1,000-grain weight. We also demonstrated that the maize (Zea mays) ZmPAR1, which is homologous to OsPAR1, is also involved in the regulation of grain size, implying a conserved role in the grain size of cereal crops. Notably, a critical SNP in the OsPAR1 coding region, causing a Glu-to-Gly substitution, is associated with grain size variation in cultivated rice. Collectively, we uncover the crucial role of OsPAR1 in grain size control, which offers a genetic basis for rice breeding in the future.
Size is a critical determinant of rice grain appearance quality and yield. In this study, we isolated a dominant mutant of rice grain size, RE10498, from an EMS-induced mutant library of ZH8015 characterized by long grai...Size is a critical determinant of rice grain appearance quality and yield. In this study, we isolated a dominant mutant of rice grain size, RE10498, from an EMS-induced mutant library of ZH8015 characterized by long grains. Using the MutMap method, we cloned the causal gene, OsGLW3, which encodes an expressed protein of unknown function. Targeted editing of position 3,119 in the OsGLW3 coding region to "A" in the wild-type ZH8015 produced positive transgenic lines whose grain size matched that of RE10498. Knocking out OsGLW3 in the wild-type ZH8015 did not alter the grain size of positive plants (KO-OsGLW3ZH), whereas knocking out OsGLW3 in the mutant RE10498 resulted in decreased grain length and increased grain width in its positive plants (KO-OsGLW3RE), reverting to a phenotype consistent with the wild type. Overexpression of the OsGLW3RE allele in ZH8015 resulted in a highly significant increase in grain length. These findings indicate that RE10498 is a gain-of-function mutant. OsGLW3RE positively regulated the level of brassinolide, which affected grain size by regulating the proliferation and extension of glume cells. Crossing ZH8015 and RE10498 with the 3-line sterile lines Fu387A (F387A), Nei5A (N5A), and Zhongxiang131A (ZX131A) showed that the hybrids F387A/RE10498, N5A/RE10498, and ZX131A/RE10498, respectively, exhibited improved rice appearance quality: longer grains with reduced chalkiness. The OsGLW3RE allele identified in this study, along with its functional marker dCAPs-G, holds significant reference value for breeding high-quality hybrid rice varieties.
Disanthus cercidifolius subsp. longipes (DCL), a rare and endangered species endemic to southern China, possesses remarkable ornamental value and thus warrants enhanced conservation and utilization. However, the lack of...Disanthus cercidifolius subsp. longipes (DCL), a rare and endangered species endemic to southern China, possesses remarkable ornamental value and thus warrants enhanced conservation and utilization. However, the lack of a reference genome has hindered research progress. Here, we report a high-quality, chromosome-scale genome assembly of DCL using PacBio HiFi and Hi-C technologies, yielding a 1.05 Gb genome with a contig N50 of 121 Mb and anchoring 99.98% of sequences to 8 chromosomes. Gene family analysis revealed that expanded gene families are mainly related to flavonoid biosynthesis and glycosyltransferase activity. DCL underwent only the whole-genome triplication event shared by core eudicots. Integrating ATAC-seq and ChIP-seq datasets, we generated a comprehensive epigenomic landscape comprising 45,359 accessible chromatin regions (ACRs). Highly expressed genes were consistently associated with a greater number of ACRs, a trend evident across both whole-genome duplication-derived gene pairs and tandem duplicates. Gene expression analysis indicated that H3K27ac, H3K9ac, and H3K4me3 may serve as critical histone marks for active regulatory elements. We further elucidated the epigenetic regulation of anthocyanin biosynthesis, mapping a molecular regulatory network. A key enhancer was involved in transcriptional regulation of Dcl09116 (chalcone isomerase) through recruitment of specific transcription factors. Collectively, this study provides comprehensive genomic and epigenomic resources for DCL, facilitating genetic improvement of its ornamental traits and advancing conservation efforts.
Little is known about how mature Arabidopsis thaliana (Arabidopsis) plants become competent to initiate Age-Related Resistance (ARR) to normally virulent Pseudomonas syringae pv tomato (Pst). RNA-Sequencing was used to i...Little is known about how mature Arabidopsis thaliana (Arabidopsis) plants become competent to initiate Age-Related Resistance (ARR) to normally virulent Pseudomonas syringae pv tomato (Pst). RNA-Sequencing was used to identify genes that contribute to ARR by comparing the response to Pst in young susceptible 3.5 weeks post germination (wpg) to mature 6 wpg ARR-competent plants. Comparative transcriptomics revealed that ARR shares many genes with Pathogen Associated Molecular Pattern (PAMP) Triggered Immunity, Effector Triggered Immunity and Systemic Acquired Resistance, providing additional evidence for the emerging idea that plant immune pathways are intertwined and interconnected. ARR-responding leaves expressed genes for the biosynthesis of N-hydroxypipecolic acid (NHP), a signaling molecule involved in Systemic Acquired Resistance (SAR) during the establishment of a primed state in systemic leaves. NHP biosynthesis mutants (ald1-1, fmo1-1) were ARR-defective and accumulated less intercellular salicylic acid (SA) compared to wild-type Col-0, indicating a requirement for functional NHP biosynthesis genes for intercellular SA accumulation during ARR. SA perception by NPR1/4 proteins was required for ARR as demonstrated by the ARR-defective phenotype of the SA-signaling mutant, npr1-1 npr4-4D. NHP biosynthesis genes were still expressed in npr1-1 npr4-4D, suggesting that similar to SAR, NHP functions upstream of NPR1-mediated signaling during ARR. Unlike ARR-incompetent plants (3, 4, 5 wpg untreated), ARR-competent plants (6 wpg untreated), expressed NHP biosynthesis genes, accumulated NHP and expressed immunity-associated cell-surface receptors RLP23 and RLP28, supporting the idea that NHP contributes to initiating a developmentally controlled ARR competent primed state in mature Arabidopsis that is similar to the primed state established in distant leaves during SAR.
Takenoya M, Shimizu T, Miyake K
… +1 more, Masuda T
Plant Physiol
· 2026 Jun · PMID 42176320
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Heme is synthesized in the plastid and metabolized to phytochromobilin (PΦB) in 2 enzymatic steps: heme oxygenase (HO) and PΦB synthase (HY2). In Arabidopsis thaliana (Arabidopsis), HO1/HY1/GUN2 predominantly functions f...Heme is synthesized in the plastid and metabolized to phytochromobilin (PΦB) in 2 enzymatic steps: heme oxygenase (HO) and PΦB synthase (HY2). In Arabidopsis thaliana (Arabidopsis), HO1/HY1/GUN2 predominantly functions for heme catabolism among the HO isoforms. Our previous study demonstrated that HO1 localization is altered in plastids or in the cytosol due to transcriptional start-site regulation. Introduction of either plastid- or cytosol-localized HO1 into HO1-deficient mutants resulted in recovery from the long hypocotyl, low pigmentation, and genomes uncoupling (gun) phenotypes, indicating the assembly of functional phytochromes (PHYs), as well as supporting the retrograde heme signaling hypothesis. To dissect the heme signaling and PHY assembly, we introduced either of the 2 types of regioselective bacterial HOs that produce biliverdin IXα (BVIXα) or BVIXβ/δ into Arabidopsis hy1-1. Gene introduction of either plastid- or cytosol-localized BVIXα-producing HO complemented the long hypocotyl, low pigmentation, and gun phenotypes of hy1-1. Interestingly, the introduction of BVIXβ/δ-producing HO, either in the plastid or in the cytosol, failed to complement the long hypocotyl and low pigmentation phenotypes, suggesting failure of functional PHY assembly. However, these lines restored the gun phenotype, thus supporting the heme signaling hypothesis. Based on the levels of complementation of the gun phenotype, we found that the expression of photosynthesis-associated nuclear genes (PhANGs) can be separated into PHY-dependent and PHY-independent groups. Our results demonstrate that heme functions as a retrograde mobile biogenic signal from plastids, which is mediated by the cytosol, to regulate the expression of PhANGs, and this regulation is distinct in its dependency on PHY.
Plant-insect coevolution has been a major driver of specialized metabolite diversification, yet the genetic basis of natural variation in defensive chemistry remains poorly understood. The wild crucifer winter cress (Bar...Plant-insect coevolution has been a major driver of specialized metabolite diversification, yet the genetic basis of natural variation in defensive chemistry remains poorly understood. The wild crucifer winter cress (Barbarea vulgaris) comprises two ecotypes, an insect-resistant G-type and a susceptible P-type, characterized by distinct triterpenoid saponin profiles. To investigate the causal relationship between saponin composition and insect feeding preference, we established a stable transformation system for B. vulgaris. P-type B. vulgaris accumulates lupeol derived saponins and expression of the G-type β-amyrin synthase gene LUP5 in the susceptible P-type conferred up to a 95% reduction in diamondback moth (Plutella xylostella) feeding, accompanied by increased accumulation of three hederagenin-derived monodesmosidic saponins. Comparison of LUP5 expression driven by its native promoter and by the constitutive 35S promoter revealed that the native promoter is activated in young leaves, but not in young developing shoots, and leads to increased hederagenin accumulation in leaves. This expression pattern reflects the coordinated expression of downstream pathway genes and prevents expression in developing shoots. Our results provide direct in planta evidence that LUP5 is a key determinant of natural variation in insect feeding preference in B. vulgaris, underscoring the pivotal role of the saponin backbone in herbivore deterrence. By linking promoter activity to metabolite structural diversity, this work provides mechanistic and conceptual insights into how plants coordinate specialized metabolism and defense.
Zhu W, Singh-Bakala H, Liu B
… +9 more, Bewg W, Bentelspacher M, Weber R, Peláez-Vico MÁ, Chen MSS, Tsai CJ, Mittler R, Yang B, Barros J
Plant Physiol
· 2026 Jun · PMID 42175571
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Cytosolic ascorbate peroxidases (APXs) have been proposed to have bifunctional 4-coumarate 3-hydroxylase (C3H) activity, linking redox regulation to lignin biosynthesis in plants. Although this dual role has been shown i...Cytosolic ascorbate peroxidases (APXs) have been proposed to have bifunctional 4-coumarate 3-hydroxylase (C3H) activity, linking redox regulation to lignin biosynthesis in plants. Although this dual role has been shown in vitro, in vivo validation remains limited. Here, we used CRISPR/Cas9 gene editing to knock out cytosolic C3H/APX genes in Brachypodium distachyon and poplar (Populus tremula × Populus alba). In Brachypodium, BdC3H/APX1 catalyzed the ascorbate-dependent hydroxylation of 4-coumarate to caffeate in vitro. Loss of BdC3H/APX1 function led to reduced lignin content, altered monomer composition, elevated H2O2 levels, and impaired growth, while double monoallelic knockouts of BdC3H/APX1 combined with a biallelic BdC3H/APX2 mutation (Bdc3h/apx1&2) exhibited severe developmental defects. Exogenous caffeate and ferulate rescued the growth and lignin phenotype of the Bdc3h/apx1 knockout mutants, whereas catalase reduced H2O2 without restoring plant growth. Similarly, CRISPR/Cas9-mediated PtC3H/APX1 knockout in poplar resulted in stunted growth and altered lignin composition, while the double Ptc3h/apx1&2 mutants were unable to regenerate from tissue culture. These results provide in vivo evidence of C3H/APX bifunctionality, suggesting that perturbed lignin biosynthesis is the primary cause of the growth defects typically observed in C3H/APX-deficient plants.
Strawberry (Fragaria × ananassa) is a globally cultivated fruit crop appreciated by consumers for its unique flavor and aroma. Major groups of volatile organic compounds that define strawberry aroma include terpenes, fa...Strawberry (Fragaria × ananassa) is a globally cultivated fruit crop appreciated by consumers for its unique flavor and aroma. Major groups of volatile organic compounds that define strawberry aroma include terpenes, fatty acid esters, furanones, and benzenoids, with complex terpene blends contributing characteristic fruity, citrus, and floral notes. Here, we report the genome-wide identification and functional analysis of the terpene synthase (TPS) family that governs terpene chemical diversity in strawberry. Mining of the allo-octoploid genome of the commercial 'Royal Royce' cultivar identified 75 TPS gene candidates. Biochemical characterization of 33 mono-, sesqui- and di-TPS enzymes via recombinant protein assays and/or microbial co-expression studies demonstrated TPS activities involved in the biosynthesis of nearly two thirds of known strawberry terpenes as well as products not previously described in octoploid strawberry. Complementary metabolomic and transcriptomic studies across a diversity panel of strawberry accessions illustrated substantial variation in the composition and abundance of more than 30 common and accession-specific terpene aroma metabolites. Analysis of two commercial accessions, 'Royal Royce' and 'Mara de Bois', with contrasting aroma profiles further revealed accession-specific alterations of terpene metabolism during fruit ripening. These findings expand our understanding of the biosynthetic genes and pathways underlying strawberry terpene metabolism and provide resources to develop breeding strategies toward improving desirable strawberry aroma traits.
Bacterial pathogens require carbohydrates from host plants to support their proliferation, in which sugar transporters play critical roles. Sucrose transporters (SUTs) execute phloem loading, while their functions in the...Bacterial pathogens require carbohydrates from host plants to support their proliferation, in which sugar transporters play critical roles. Sucrose transporters (SUTs) execute phloem loading, while their functions in the plant-pathogen interaction are largely unknown. Here, we pinpoint the role of SlSUT2 during Pseudomonas syringae pv tomato (Pst) DC3000 infection of tomato plants. SlSUT2 transcription is positively correlated with bacterial proliferation during Pst DC3000 infection. Knocking out SlSUT2 impairs plant growth and enhances resistance to the pathogen. The slsut2 mutant has reduced sugar contents in both the apoplast and phloem and more starch accumulation in photosynthetic leaf cells. Pst DC3000 induces the expression of the transcription factor SlMYC2, which binds to the SlSUT2 promoter and activates its expression. SlSUT2 interacts with the main phloem loading transporter SlSUT1 and boosts its accumulation on the plasma membrane, suggesting that defective sugar transport in the slsut2 mutant is possibly due to reduced SlSUT1 accumulation. Knocking out SlSUT1 increases the resistance against Pst DC3000 and reduces sugar content in both the apoplast and phloem. Transcriptome analysis reveals that the slsut2 mutant has enhanced SA biosynthesis and signaling. Taken together, our study demonstrates that Pst DC3000 utilizes tomato SlSUT2 to trigger sugar transport and repress SA-mediated resistance to support pathogen growth in plants, suggesting SlSUT2 as a potential target for disease control and resistance breeding.
The phytohormone cytokinin has diverse functions in plants, most of which have been primarily elucidated in the eudicotyledonous plant Arabidopsis (Arabidopsis thaliana). To further characterize the role of cytokinin in...The phytohormone cytokinin has diverse functions in plants, most of which have been primarily elucidated in the eudicotyledonous plant Arabidopsis (Arabidopsis thaliana). To further characterize the role of cytokinin in a monocot, we used CRISPR-Cas9 to generate rice (Oryza sativa) lines with frameshift alleles in genes encoding the 4 putative histidine kinase (HK) cytokinin receptors, resulting in all single and multiple mutant combinations. The hk3,4,5,6 quadruple mutant transcriptome was completely insensitive to cytokinin, suggesting that all transcription regulated through cytokinin signaling acts through these 4 HKs, despite the presence of a putative cytokinin-binding CHASE-containing Ser/Thr kinase in rice. Rice HKs functioned partially redundantly in diverse aspects of growth and development, with different HKs playing more prominent roles in various processes. This functional specialization was most striking in panicle architecture, where HK4 promoted fertility, HK3 and HK5 drove secondary branch formation, and HK6 drove primary branch formation. Furthermore, unlike Arabidopsis, cytokinin signaling was essential for shoot formation in rice, as the hk3,4,5,6 quadruple mutant was shootless, likely due to a failure to develop a shoot apical meristem. The necessity for shoot formation is consistent with one of the first roles of cytokinin uncovered by Skoog and Miller in the 1950s. Overall, this work elucidates the many roles of cytokinin perception throughout the lifespan of rice, some of which diverge from those known in Arabidopsis, highlighting the need to study alternative model systems.
Bud dormancy release in woody plants is crucial for survival, regrowth, flowering, and fruiting. Tree peony (Paeonia suffruticosa), an important ornamental and economic plant, undergoes bud endodormancy in winter, and su...Bud dormancy release in woody plants is crucial for survival, regrowth, flowering, and fruiting. Tree peony (Paeonia suffruticosa), an important ornamental and economic plant, undergoes bud endodormancy in winter, and sufficient chilling duration and exogenous gibberellins (GAs) can effectively break the dormancy. However, the epigenetic regulation mechanism remains poorly understood. Here, immunoblotting revealed that H3K4me3, but not H3K4me1 or H3K4me2, was associated with chilling- and GA3-induced dormancy release. Chromatin immunoprecipitation sequencing (ChIP-seq) combined with RNA-seq results revealed that H3K4me3 enriched near transcription start sites (TSS). H3K4me3 enrichment genes (HEGs) and differentially expressed genes (DEGs) were commonly enriched in KEGG pathways, such as plant hormone signal transduction and MAPK signaling. The expression patterns of these marker genes, such as EARLY BUD-BREAK 3 (PsEBB3), CYCLIND3.1 (PsCYCD3.1), CYCLIND3.3 (PsCYCD3.3), and β-1,3-glucanase 6 (PsBG6), were correlated with their H3K4me3 enrichment and were validated by chromatin immunoprecipitation-quantitative PCR (ChIP-qPCR). Four COMPASS-like component homologs and one histone methyltransferase were screened; among them, PsWDR5a, PsRBL, PsASH2R, and PsATX1 were up-regulated by prolonged chilling and GA3 treatments. Yeast two-hybrid (Y2H), yeast three-hybrid (Y3H), luciferase complementation (LCA), and co-immunoprecipitation (Co-IP) analyses revealed that PsRBL interacts with PsWDR5a and PsASH2R as a bridge. PsATX1 was confirmed as an H3K4me3 methyltransferase and interacted with PsWDR5a and PsRBL to form the PsATX1-COMPASS-like complex using Y2H, LCA, and Co-IP assays. Functional analyses showed that PsWDR5a, PsRBL, PsASH2R, and PsATX1 significantly promoted budburst by elevating genomic H3K4me3 levels. Our findings provide insights into the epigenetic regulation of dormancy transitions in woody perennials.