Shoot branching is an important crop agronomic trait that directly affects plant architecture and crop productivity. Although phytochrome B (phyB), BRANCHED1 (BRC1), and abscisic acid (ABA) mediate axillary bud outgrowth, it is unknown if there is any integrating factor among them in the Plantae. We report that mutation of CsphyB or inactivation of CsphyB by shade inhibits lateral bud outgrowth in cucumber. Cucumber PHYTOCHROME INTERACTING FACTOR 4 (CsPIF4) interacts with CsphyB and directly binds to the promoter of CsBRC1 to activate CsBRC1 expression. CsBRC1 also directly promotes the expression of ABA biosynthesis gene 9-CIS-EPOXICAROTENOID DIOXIGENASE 3 (CsNCED3). Functional disruption of CsPIF4 decreased expression of CsBRC1 and CsNCED3, reduced ABA accumulation, and increased bud outgrowth in cucumber. Csnced3 mutants had reduced ABA levels and increased lateral bud outgrowth. These results suggest that a regulatory network involving CsphyB-CsPIF4-CsBRC1 exists that integrates light signaling and ABA biosynthesis to modulate bud outgrowth. This provides a strategy to manipulate branch numbers in crop breeding to realize ideal branching characteristics to maximize yield.

Water-soluble protein (WSP) content determines soybean nutritional value and processing efficiency, yet its genetic and molecular regulation remains poorly understood. Here, we identified Glycine max gibberellic acid-stimulated Arabidopsis 12 (GmGASA12), encoding a gibberellin-regulated protein, as a major quantitative trait locus (QTL) governing WSP, through genome-wide association studies across five environments. Knockout of GmGASA12 resulted in 28.7% higher WSP content, 27.6% enlarged seed cells, and 20% yield increase, while overexpression suppressed these traits. Hormonal profiling revealed that GmGASA12 knockout elevates gibberellin, auxin, and abscisic acid but reduces cytokinin, driving cell expansion and protein body accumulation. GmGASA12 cooperatively regulates the biosynthesis of β-conglycinin and glycinin, the core storage proteins in soybean seeds, through its interaction with GmCG-6. Transcriptomics linked GmGASA12 to nitrogen metabolism and hormone signaling, with knockout upregulating amino acid transporters (GmAAP3/6/27) and storage protein genes (GmCG-1–6). Evolutionary analyses demonstrated strong selection for elite GmGASA12 haplotypes during domestication, with 94% of cultivars harboring favorable alleles. Our findings establish GmGASA12 as a molecular hub integrating hormonal dynamics and protein interaction to enhance soybean quality and yield, offering a pivotal target for breeding nutrient-dense varieties.

Potassium (K⁺), an essential macronutrient, strongly influences myriad fundamental processes, while its deficiency inhibits plant growth. Jasmonic acid (JA) regulates plant growth; however, its role in plant growth inhibition under K⁺ deficiency remains nebulous. Herein, we determined that JA significantly inhibits low K⁺ tolerance and K⁺ uptake in tomato. Methyl jasmonate treatment induced the expression of SlMYC3-like under low K⁺ stress, which bound the promoters of the genes that encode KT/KUP/HAK-type transporter (SlHAK5) and voltage-gated K⁺ channel (SlLKT1) and inhibited their expression. Knockdown of SlMYC3-like enhanced low K⁺ stress tolerance and decreased JA responses, while its overexpression led to low K⁺ stress sensitivity and promoted jasmonate responses in tomato. In addition, jasmonate ZIM-domain transcriptional repressor 2/3 (SlJAZ2/3) interacted with SlMYC3-like; this interaction decreased DNA-binding activity of SlMYC3-like. SlMYC3-like promoted SlJAZ2/3 expression, forming a negative feedback circuit in JA signaling. Silencing SlJAZ2/3 increased plant susceptibility to low K⁺ stress. Our findings demonstrate the involvement of the JA–SlJAZ2/3–SlMYC3-like module in K⁺ uptake and plant growth in tomato under low K⁺ stress, providing novel insights into the regulation of plant growth and K⁺ uptake.

The calcineurin B-like protein (CBL)-CBL-interacting protein kinase (CIPK) Ca²⁺ sensors play crucial roles in the plant's response to drought stress. However, there have been few reports on the synergistic regulation of drought stress by CBL-CIPK and abscisic acid (ABA) core signaling components. In this study, we discovered that ZmCIPK33 positively regulates drought resistance in maize. ZmCIPK33 physically interacts with and is enhanced by phosphorylation from ZmSnRK2.10. Drought stress can activate ZmCIPK33, which is partially dependent on ZmSnRK2.10. ZmCIPK33 in combination with ZmSnRK2.10 can activate the slow anion channel ZmSLAC1 in Xenopus laevis oocytes independently of CBLs, whereas ZmCIPK33 or ZmSnRK2.10 alone is unable to do so. Furthermore, ZmCIPK33 phosphorylates ZmPP2C11 at Ser60, which leads to a reduction in the interaction between ZmPP2C11 and ZmEAR1 (the ortholog of Arabidopsis Enhancer of ABA co-Receptor 1) and weakens the phosphatase activity of ZmPP2C11, consequently, enhancing the activity of ZmSnRK2.10 in an in vitro assay and in the in-gel assay of the zmcipk33 mutant. Our findings provide novel insights into the molecular mechanisms underlying the reciprocal enhancement of Ca²⁺ and ABA signaling under drought stress in maize.

Radish (Raphanus sativus L.) is a globally important root vegetable crop known for its diverse varieties and unique taproot characteristics. The LBD (LATERAL ORGAN BOUNDARIES DOMAIN) gene family, specific to plants, plays a pivotal role in the development of lateral plant organs. Nonetheless, the precise biological functions and molecular regulatory mechanisms of LBD genes in radish taproot development remain largely unexplored. In this study, the RsLBD3 gene was identified as a potential candidate affecting taproot size in radish through a genome-wide association study. Further investigation revealed two insertions in the C-terminal region of RsLBD3, with insertion³⁶³ notably enhancing the transcriptional activation capability of RsLBD3. It was observed that radish taproots with RsLBD3^Ins-363 haplotype displayed significantly greater length and weight compared to those with RsLBD3^Del-363 haplotype. RNA in situ hybridization and reverse transcription quantitative polymerase chain reaction analysis revealed that the RsLBD3 gene exhibits high expression level in the vascular cambium and is induced by cytokinin treatment. Silencing the RsLBD3 gene resulted in the inhibition of vascular cambium activity in the taproot, thereby impeding thickening. Exogenous cytokinin treatment could partially rescue the small-taproot phenotypes caused by RsLBD3 silencing. Moreover, RsARF5 (AUXIN RESPONSE FACTOR 5), RsRR7b (RESPONSE REGULATOR 7), and RsCYCD3-1 (CYCLIN D3;1) were identified as target genes of RsLBD3. Notably, RsARF5 was found to directly regulate the expression of RsWOX4 (WUSCHEL-RELATED HOMEOBOX 4). Additionally, biochemical analysis demonstrated that RsTCP14 interacts with RsLBD3, contributing to the binding of RsLBD3 to its target genes. Collectively, these findings contribute to a better understanding of the regulatory mechanisms underlying taproot morphogenesis, and provide novel allelic variations for the genetic enhancement of taproot shape traits in radish.

Jasmonic acid (JA) and gibberellin (GA) coordinate many aspects of plant growth and development, including anthocyanin biosynthesis. However, the crossover points of JA and GA signals and the pathways through which they interact to regulate anthocyanin biosynthesis are poorly understood. Here, we investigated the molecular mechanism by which the zinc finger protein (ZFP) transcription factor Malus domestica ZFP7 (MdZFP7) regulates anthocyanin biosynthesis by integrating JA and GA signals at the transcriptional and post-translational levels. MdZFP7 is a positive regulator of anthocyanin biosynthesis, which fulfills its role by directly activating the expression of MdMYB1 and enhancing the transcriptional activation of MdWRKY6 on the target genes MdDFR and MdUF3GT. MdZFP7 integrates JA and GA signals by interacting with the JA repressor apple JASMONATE ZIM-DOMAIN2 (MdJAZ2) and the GA repressor apple REPRESSOR-of-ga1-3-like 3a (MdRGL3a). MdJAZ2 weakens the transcriptional activation of MdMYB1 by MdZFP7 and disrupts the MdZFP7–MdWRKY6 interaction, thereby reducing the anthocyanin biosynthesis promoted by MdZFP7. MdRGL3a contributes to the stimulation of anthocyanin biosynthesis by MdZFP7 by sequestering MdJAZ2 from the MdJAZ2–MdZFP7 complex. The E3 ubiquitin ligase apple BOI-related E3 ubiquitin-protein ligase 3 (MdBRG3), which is antagonistically regulated by JA and GA, targets the ubiquitination degradation of MdZFP7. The MdBRG3-MdZFP7 module moves the crosstalk of JA and GA signals from the realm of transcriptional regulation and into the protein post-translational modification. In conclusion, this study not only elucidates the node-role of MdZFP7 in the integration of JA and GA signals, but also describes the transcriptional and post-translational regulatory network of anthocyanin biosynthesis with MdZFP7 as the hub.

Besides playing a crucial role in plant immunity via the nonexpressor of pathogenesis-related (NPR) proteins, increasing evidence shows that salicylic acid (SA) can also regulate plant root growth. However, the transcriptional regulatory network controlling this SA response in plant roots is still unclear. Here, we found that NPR1 and WRKY45, the central regulators of SA response in rice leaves, control only a reduced sector of the root SA signaling network. We demonstrated that SA attenuates root growth via a novel NPR1/WRKY45-independent pathway. Furthermore, using regulatory network analysis and mutant characterization, we identified a set of new NPR1/WRKY45-independent regulators that conservedly modulate the root development and root-associated microbiota composition in both Oryza sativa (monocot) and Arabidopsis thaliana (dicot) in response to SA. Our results established the SA signaling as a central element regulating plant root functions under ecologically relevant conditions. These results provide new insights to understand how regulatory networks control plant responses to abiotic and biotic stresses.

Stomata are epidermal pores that are essential for water evaporation and gas exchange in plants. Stomatal development is orchestrated by intrinsic developmental programs, hormonal controls, and environmental cues. The steroid hormone brassinosteroid (BR) inhibits stomatal lineage progression by regulating BIN2 and BSL proteins in leaves. Notably, BR is known to promote stomatal development in hypocotyls as opposed to leaves; however, its molecular mechanism remains elusive. Here, we show that BR signaling has a dual regulatory role in controlling stomatal development in Arabidopsis hypocotyls. We found that brassinolide (BL; the most active BR) regulates stomatal development differently in a concentration-dependent manner. At low and moderate concentrations, BL promoted stomatal formation by upregulating the expression of SPEECHLESS (SPCH) and its target genes independently of BIN2 regulation. In contrast, high concentrations of BL and bikinin, which is a specific inhibitor of BIN2 and its homologs, significantly reduced stomatal formation. Genetic analyses revealed that BIN2 regulates stomatal development in hypocotyls through molecular mechanisms distinct from the regulatory mechanism of the cotyledons. In hypocotyls, BIN2 promoted stomatal development by inactivating BZR1, which suppresses the expression of SPCH and its target genes. Taken together, our results suggest that BR precisely coordinates the stomatal development of hypocotyls using an antagonistic control of SPCH expression via BZR1-dependent and BZR1-independent transcriptional regulation.

The phytohormone jasmonates (JAs) regulate plant growth and defense responses. The reproductive organs of flowers are devastated by insect herbivores. However, the molecular mechanisms of floral defense remain largely unknown. Here, we found that the Arabidopsis JA receptor CORONATINE INSENSITIVE1 (COI1) and its substrates JA ZIM-domain (JAZ) repressors, and the mediator subunit MEDIATOR25-based MED25–MYC–MYB (MMM) complexes, including MYC2/3/4/5 and MYB28/29/76, mediated floral defense against the insects Helicoverpa armigera, Spodoptera exigua, and Spodoptera frugiperda. The flower-specific IIIa bHLH factors ABORTED MICROSPORES (AMS) and DYSFUNCTIONAL TAPETUM 1 (DYT1) were JAZ-interaction proteins. They interacted with members of the MMM complexes, inhibited the transcriptional activity of MYC2 and MYB28, and repressed floral defense against insects. AMS and DYT1 recruited the flower-specific MYB21/24, and these MYBs interacted with members of MMM complexes, inhibited the MYC2–MYB28 function, and suppressed floral defense against insects. Our study revealed that the JA–COI1–JAZ–MMM pathway mediated flower defense, and the AMS/DYT1–MYB21/24 module antagonized the MMM complexes to repress floral defense against insects.

Plant growth is determined by the production of cells and initiation of new organs. Exploring genes that control cell number and cell size is of great significance for understanding plant growth regulation. In this study, we characterized two wheat mutants, ah and dl, with abnormal growth. The ah mutant is a naturally occurring variant characterized by severe dwarfism, increased tiller number, and reduced grain length, while the dl mutant is derived from an ethyl methane sulfonate (EMS)-mutagenized population and exhibits smaller grain size and slightly reduced plant height. Cytological analyses revealed abnormal cell number, cell morphology and arrangement in the stems and leaves of the ah mutant, along with reduced cell length in the grains of the dl mutant. Map-based cloning identified that both mutants carry mutations in the same gene TavWA1-7D, which encodes a protein with a von Willebrand factor A (vWA) domain. The ah mutant harbors a 174-bp insertion in the 1,402-bp coding sequence (CDS) of TavWA1-7D, causing premature termination of protein translation, while the dl mutant contains a Glu420Lys substitution. Mimicking the TavWA1-7D^ah through clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated nuclease 9-mediated genome editing leads to a severe dwarfism phenotype. The C-terminus of the protein is crucial for its correct subcellular localization and interaction, supporting its critical role for TavWA1-7D function. Proteomic analysis showed that the dwarf phenotype of the ah mutant is associated with impaired photosynthesis, ribosome function, and nucleosome formation. Additionally, TavWA1-7D interacts with an E3 ligase, TaVIP1-3B, the expression levels of which are elevated in both mutants. Overexpression and knockout studies of TaVIP1-3B demonstrated its negative regulatory role in cell length and grain size. Together, our findings suggest that TavWA1-7D plays a vital role in regulating wheat growth and yield-related traits, with the dl mutant's short grain phenotype being associated with TaVIP1-3B expression levels.

WRKY transcription factors play key roles in plant resistance to various stresses, but their roles in fruit ripening remain largely unknown. Here, we report a WRKY gene PpWRKY14 involved in the regulation of fruit ripening in peach. The expression of PpWRKY14 showed an increasing trend throughout fruit development. PpWRKY14 was a target gene of PpNAC1, a master regulator of peach fruit ripening. PpWRKY14 could directly bind to the promoters of PpACS1 and PpACO1 to induce their expression, and this induction was greatly enhanced when PpWRKY14 formed a dimer with PpNAC1. However, the transcription of PpNAC1 could be directly suppressed by two EIN3/EIL1 genes, PpEIL2 and PpEIL3. The PpEIL2/3 genes were highly expressed at the early stages of fruit development, but their expression was programmed to decrease significantly during the ripening stage, thus derepressing the expression of PpNAC1. These results suggested a PpEIL2/3–PpNAC1–PpWRKY14 module that regulates fruit ripening by modulating ethylene production in peach. Our results provided an insight into the regulatory roles of EIN3/EIL1 and WRKY genes in fruit ripening.

The plant hormone jasmonate (JA) regulates plant growth and immunity by orchestrating a genome-wide transcriptional reprogramming. In the resting stage, JASMONATE-ZIM DOMAIN (JAZ) proteins act as main repressors to regulate the expression of JA-responsive genes in the JA signaling pathway. However, the mechanisms underlying de-repression of JA-responsive genes in response to JA treatment remain elusive. Here, we report two nuclear factor Y transcription factors NF-YB2 and NF-YB3 (thereafter YB2 and YB3) play key roles in such de-repression in Arabidopsis. YB2 and YB3 function redundantly and positively regulate plant resistance against the necrotrophic pathogen Botrytis cinerea, which are specially required for transcriptional activation of a set of JA-responsive genes following inoculation. Furthermore, YB2 and YB3 modulated their expression through direct occupancy and interaction with histone demethylase Ref6 to remove repressive histone modifications. Moreover, YB2 and YB3 physically interacted with JAZ repressors and negatively modulated their abundance, which in turn attenuated the inhibition of JAZ proteins on the transcription of JA-responsive genes, thereby activating JA response and promoting disease resistance. Overall, our study reveals the positive regulators of YB2 and YB3 in JA signaling by positively regulating transcription of JA-responsive genes and negatively modulating the abundance of JAZ proteins.

Most mechanistic details of chronologically ordered regulation of leaf senescence are unknown. Regulatory networks centered on AtWRKY53 are crucial for orchestrating and integrating various senescence-related signals. Notably, AtWRKY53 binds to its own promoter and represses transcription of AtWRKY53, but the biological significance and mechanism underlying this self-repression remain unclear. In this study, we identified the VQ motif-containing protein AtVQ25 as a cooperator of AtWRKY53. The expression level of AtVQ25 peaked at mature stage and was specifically repressed after the onset of leaf senescence. AtVQ25-overexpressing plants and atvq25 mutants displayed precocious and delayed leaf senescence, respectively. Importantly, we identified AtWRKY53 as an interacting partner of AtVQ25. We determined that interaction between AtVQ25 and AtWRKY53 prevented AtWRKY53 from binding to W-box elements on the AtWRKY53 promoter and thus counteracted the self-repression of AtWRKY53. In addition, our RNA-sequencing data revealed that the AtVQ25-AtWRKY53 module is related to the salicylic acid (SA) pathway. Precocious leaf senescence and SA-induced leaf senescence in AtVQ25-overexpressing lines were inhibited by an SA pathway mutant, atsid2, and NahG transgenic plants; AtVQ25-overexpressing/atwrky53 plants were also insensitive to SA-induced leaf senescence. Collectively, we demonstrated that AtVQ25 directly attenuates the self-repression of AtWRKY53 during the onset of leaf senescence, which is substantially helpful for understanding the timing of leaf senescence onset modulated by AtWRKY53.

Panicle exsertion is one of the crucial agronomic traits in rice (Oryza sativa). Shortening of panicle exsertion often leads to panicle enclosure and severely reduces seed production. Gibberellin (GA) plays important roles in regulating panicle exsertion. However, the underlying mechanism and the relative regulatory network remain elusive. Here, we characterized the oswrky78 mutant showing severe panicle enclosure, and found that the defect of oswrky78 is caused by decreased bioactive GA contents. Biochemical analysis demonstrates that OsWRKY78 can directly activate GA biosynthesis and indirectly suppress GA metabolism. Moreover, we found OsWRKY78 can interact with and be phosphorylated by mitogen-activated protein kinase (MAPK) kinase OsMAPK6, and this phosphorylation can enhance OsWRKY78 stability and is necessary for its biological function. Taken together, these results not only reveal the critical function of OsWRKY78, but also reveal its mechanism via mediating crosstalk between MAPK and the GA signaling pathway in regulating panicle exsertion.

Anthocyanins are secondary metabolites induced by environmental stimuli and developmental signals. The positive regulators of anthocyanin biosynthesis have been reported, whereas the anthocyanin repressors have been neglected. Although the signal transduction pathways of gibberellin (GA) and jasmonic acid (JA) and their regulation of anthocyanin biosynthesis have been investigated, the cross-talk between GA and JA and the antagonistic mechanism of regulating anthocyanin biosynthesis remain to be investigated. In this study, we identified the anthocyanin repressor MdbHLH162 in apple and revealed its molecular mechanism of regulating anthocyanin biosynthesis by integrating the GA and JA signals. MdbHLH162 exerted passive repression by interacting with MdbHLH3 and MdbHLH33, which are two recognized positive regulators of anthocyanin biosynthesis. MdbHLH162 negatively regulated anthocyanin biosynthesis by disrupting the formation of the anthocyanin-activated MdMYB1-MdbHLH3/33 complexes and weakening transcriptional activation of the anthocyanin biosynthetic genes MdDFR and MdUF3GT by MdbHLH3 and MdbHLH33. The GA repressor MdRGL2a antagonized MdbHLH162-mediated inhibition of anthocyanins by sequestering MdbHLH162 from the MdbHLH162-MdbHLH3/33 complex. The JA repressors MdJAZ1 and MdJAZ2 interfered with the antagonistic regulation of MdbHLH162 by MdRGL2a by titrating the formation of the MdRGL2a-MdbHLH162 complex. Our findings reveal that MdbHLH162 integrates the GA and JA signals to negatively regulate anthocyanin biosynthesis. This study provides new information for discovering more anthocyanin biosynthesis repressors and explores the cross-talk between hormone signals.

Secondary vascular tissue (SVT) development and regeneration are regulated by phytohormones. In this study, we used an in vitro SVT regeneration system to demonstrate that gibberellin (GA) treatment significantly promotes auxin-induced cambium reestablishment. Altering GA content by overexpressing or knocking down ent-kaurene synthase (KS) affected secondary growth and SVT regeneration in poplar. The poplar DELLA gene GIBBERELLIC ACID INSENSITIVE (PtoGAI) is expressed in a specific pattern during secondary growth and cambium regeneration after girdling. Overexpression of PtoGAI disrupted poplar growth and inhibited cambium regeneration, and the inhibition of cambium regeneration could be partially restored by GA application. Further analysis of the PtaDR5:GUS transgenic plants, the localization of PIN-FORMED 1 (PIN1) and the expression of auxin-related genes found that an additional GA treatment could enhance the auxin response as well as the expression of PIN1, which mediates auxin transport during SVT regeneration. Taken together, these findings suggest that GA promotes cambium regeneration by stimulating auxin signal transduction.