Molecular Plant

Heavy metal-associated isoprenylated plant proteins (HIPPs) are encoded by large gene families, which have diversified specifically in vascular plants. Their physiological functions and molecular mode of activity are currently largely unknown. In this study, we characterize a group of phylogenetically closely related genes HIPP32, HIPP33, and HIPP34 in Arabidopsis thaliana, revealing their essential roles in controlling diverse developmental pathways. Through comprehensive genetic analyses, we demonstrate that these genes exhibit partially overlapping pleiotropic functions, influencing multiple aspects of plant growth such as embryogenesis, maintenance of apical meristems, root architecture, shoot branching, leaf morphogenesis and floral organ formation. Transcriptomic profiling of hipp mutants identified significant deregulation in several regulatory pathways involved in plant hormone responses, with a specific impact on auxin signaling processes. Interestingly, we show that the analyzed HIPP proteins localize very specifically to plasmodesmata, suggesting their potential function in regulating intercellular communication in shaping plant development.

Stem development is crucial in plant vertical architecture and overall crop improvement. The molecular mechanisms underlying the initiation and circadian growth of stem remain still enigmatic. Here, we demonstrate that APETALA1 (AP1) spatially initiates circadian stem growth through a noncell-autonomous role of TIMING OF CAB EXPRESSION1 (TOC1) emanating from the floral meristem to inflorescence meristem in Arabidopsis thaliana. Mechanistically, AP1 interacts with and recruits CIRCADIAN CLOCK-ASSOCIATED1 (CCA1) to associate TOC1. Dynamic formation of the AP1-CCA1 complex is crucial for maintaining the circadian rhythmicity and expression of TOC1. Tissue-specific expression of TOC1 in floral meristems rescues the developmental defects of toc1 mutants at both functional and transcriptional levels. Furthermore, TOC1 directly activates the circadian expression of pectin methylesterase gene family, which is critical for its role in stem development. AP1 homologs regulate circadian stem elongation and plant height in wheat and rice, underscoring the conserved mechanism across flowering plants. Our findings uncovered a spatiotemporal regulatory cascade gating stem development and identified candidate genes for crop improvement.

Chromosome number variation and structural reorganization are key drivers of plant evolution, yet their genomic basis remains unclear due to incomplete representation of repetitive regions in existing assemblies. The Linum genus exhibits exceptional karyotypic diversity (n = 7-43), providing a powerful system to investigate chromosome evolution. Here, we generated near telomere-to-telomere (T2T) genome assemblies for four species, including cultivated flax (L. usitatissimum cv. CDC Bethune; n = 15), its wild progenitor (L. bienne; n = 15), and two related species (L. decumbens and L. grandiflorum; n = 8). Together with published genomes of L. lewisii (n = 9) and L. tenue (n = 10), these enabled reconstruction of chromosome evolution across six lineages. Phylogenomic analyses revealed a shared ancestral whole-genome duplication (WGD) associated with the n = 9 karyotype, followed by lineage-specific WGDs and divergent diploidization. The transition from n = 8 to the derived n = 15 flax lineage not only occurred without chromosome length expansion, but also with genome size reduction, indicating extensive internal restructuring. Comparative analyses showed that this restructuring was associated with lineage-specific expansion of a single DNA transposon family (TE_00003234; hAT), which is highly enriched in expansive pericentromeric regions that are characterized by low gene density and nucleotide diversity, suppressed recombination, segregation distortion, and extensive synteny disruption, unlike the LTR retrotransposon-rich pericentromeres typical of most plant genomes. These findings support a model in which lineage-specific DNA transposon expansion is associated with remodeling of pericentromeric architecture and large-scale chromosome restructuring following polyploidization.

High humidity greatly influences plant growth and development and triggers adaptive physiological responses such as leaf hyponasty (elongation of leaf petiole and upward leaf movement). A recent study identified Cyclic Nucleotide-Gated Ion Channels 2 and 4 (CNGC2/4)-mediated Ca2+ influx and Calmodulin Binding Transcription Activators 2 and 3 (CAMTA2/3)-mediated transcription as essential for high humidity response in Arabidopsis, but the upstream regulators that control these pathways remain unknown. Here, we show that the receptor-like kinase FERONIA and its co-receptor LORELEI-LIKE GPI-ANCHORED PROTEIN1 (LLG1) are required for a large portion of high humidity-associated Arabidopsis transcriptomic changes, including CNGC2, CAMTA-regulated genes, and cell wall remodeling genes, and for high humidity-induced leaf hyponasty. High humidity triggers a previously uncharacterized petiole-localized Ca2+ waves that precede hyponastic leaf movement. The petiole-localized Ca2+ signals were significantly altered in the fer-4 mutant. Thus, FERONIA is a key regulator of plant responses to extracellular high humidity. Highlights FERONIA plays a prominent role in transcriptomic responses to high humidity FERONIA is required for high humidity-induced leaf hyponasty High humidity induces petiole calcium waves FERONIA is required for normal petiole calcium waves in response to high humidity

Photomorphogenesis, the light-driven development of seedlings, is governed by a complex network of transcription factors and circadian regulators. While the TIMING OF CAB EXPRESSION 1 (TOC1) is known to link circadian rhythms with light-responsive growth, the mechanisms fine-tuning its activity remain poorly understood. Here, we identify PHOSPHATE 1 HOMOLOG 2 (PHO1;H2) as a novel negative regulator of seedling photomorphogenesis in Arabidopsis. Loss-of-function pho1;h2 mutants exhibit hypersensitivity to light, characterized by markedly shorter hypocotyls and increased photopigment accumulation, whereas overexpression lines display reduced photomorphogenic response. We demonstrate that the N-terminal SPX domain of PHO1;H2 is both necessary and sufficient to repress seedling photomorphogenic growth. Mechanistically, in vitro and in vivo interaction assays reveal that the SPX domain physically binds and sequesters TOC1, inhibiting its regulatory function. This PHO1;H2-mediated sequestration of TOC1 alleviates the repression of PHYTOCHROME INTERACTING FACTOR 4 (PIF4), thereby promoting the expression of downstream genes involved in cell elongation and hormone signaling. Collectively, our findings reveal a competitive binding mechanism by which PHO1;H2 modulates the TOC1-PIF4 signaling axis, providing a crucial checkpoint for seedling growth in dynamic light environments.

Sugar is both an essential energy source and the major substrate for cell wall biosynthesis during plant growth, yet how growth-promoting hormones regulate sugar synthesis remains unclear. Here, we show that the brassinosteroids (BRs) promote gluconeogenic and photosynthetic sugar synthesis by activating phosphoenolpyruvate carboxykinase (PCK), which catalyzes the conversion of oxaloacetate to phosphoenolpyruvate, a central step in primary metabolism. Arabidopsis BR-deficient mutants display reduced PCK1 activity and elevated phosphorylation at conserved Ser-62 and Thr-66 residues. BR treatment induces PCK1 dephosphorylation and activation, whereas the GSK3-like kinase BIN2 phosphorylates these sites, altering quaternary structure and inhibiting PCK1. Phospho-blocking mutations of Ser-62/Thr-66 confer BR-independent PCK1 activity and enhance seedling growth, while phosphomimetic mutations reduce PCK1 activity and impair seedling growth and establishment. BR also promotes PCK dephosphorylation and activation in photosynthetic leaves of maize and sorghum. Our study demonstrates that BR regulates primary metabolism via GSK3/BIN2-mediated phosphorylation of PCK, thereby promoting gluconeogenesis and photosynthesis.

In nature, plants are subjected to multiple environmental stress factors simultaneously or sequentially. Recent studies revealed that when three or more stress factors impact a plant simultaneously (termed multifactorial stress combination; MFSC), plant survival declines, even if the intensity of each individual stress involved in the MFSC is low. We previously identified RAP2.3 as a key transcription factor (TF) required for Arabidopsis thaliana survival, specifically under a MFSC of salt+excess light+heat stress (i.e., S+EL+HS). Here we report that RAP2.3 is required for the expression of SIGMA3, a nuclear-encoded factor that directs plastid RNA polymerase to specific plastid promoters, and MYB51, a key stress response TF involved in glucosinolate metabolism and oxidative stress responses, specifically during a MFSC of S+EL+HS. Like rap2.3 mutants, myb51 and sig3 mutants display significantly low survival rate specifically under the MFSC of S+EL+HS. Based on MYB51 gene regulatory network analysis and characterization of jasmonic acid (JA) mutants, we further reveal that suppression of JA signaling could play an important role in promoting plant survival under conditions of S+EL+HS. Our findings uncover an additional layer of the response of plants to MFSC, as well as identify potential targets for breeding crops with enhanced tolerance to climate change.

Photomorphogenesis allows plants to adjust growth to ambient light conditions and relies on protein quality control to ensure the timely turnover of signaling components. The conserved AAA+ ATPase CDC48, along with its cofactors NPL4 and UFD1, is a crucial regulator of proteasomal degradation. While well characterized in other organisms, its role in plant development remains largely unexplored. Here, we show that CDC48 is required for blue light-mediated photomorphogenesis in Arabidopsis. Under blue light, CDC48A accumulates at the plasma membrane and in the nucleus, and cdc48a mutants fail to repress hypocotyl elongation properly. Similar phenotypes are observed upon inhibition of CDC48 or in npl4 and ufd1 mutants. Genetic and biochemical analyses further reveal that CDC48A negatively regulates gibberellin (GA) signaling. Consistently, UFD1 directly interacts with the GA receptor GID1 to promote its degradation. Together, these findings demonstrate that CDC48A integrates light and hormonal cues through protein homeostasis to regulate photomorphogenic development.

Plants, as sessile organisms, must continually adapt to fluctuating temperatures to ensure survival. The plasma membrane-localized receptor-like kinase FERONIA (FER) coordinates diverse physiological processes and responses to various biotic and abiotic stresses. However, the role of FER in plant adaptation to elevated temperatures remains largely unexplored. Here, we report that FER is indispensable for plant thermotolerance. We found that fer loss-of-function mutants exhibit impaired thermomorphogenic growth and are hypersensitive to mild heat stress, displaying extensive oxidative stress-mediated cell death at elevated temperatures. Combined genetic and molecular analyses revealed that these temperature-sensitive defects in fer mutants are caused by an overaccumulation of jasmonic acid (JA), which subsequently triggers excessive production of reactive oxygen species. Furthermore, we show that this aberrant JA accumulation and oxidative stress are attributable to impaired FER-mediated regulation of turgor-dependent cell wall tensile stress. Taken together, our results suggest that FER-mediated cell wall tensile stress regulation serves as a critical mechanism to prevent aberrant JA accumulation and oxidative stress at elevated temperatures, thereby enabling plants to adapt to and survive under high-temperature conditions.

Oryza granulata Nees et Arn. ex Watt, a diploid wild rice (GG genome), possesses exceptional shade tolerance and is a key genetic resource for rice improvement. However, previous genome assemblies lacked continuity and completeness. Here we present a chromosome-scale reference genome of O. granulata using PacBio SMRT (113x), Hi-C (95x), and Illumina sequencing. The final assembly is ~764.24 Mb, with a scaffold N50 of ~59.32 Mb, and ~96.47% of the sequence anchored to 12 chromosomes. BUSCO completeness is ~98.6%. We annotated ~42,064 protein-coding genes, of which ~95.39% were functionally annotated, along with ~73.46% repetitive elements. The genome assembly and raw sequencing data are available at NGDC (PRJCA061980), NGDC GSA (CRA068332), and NGDC GWH (GWHISVE00000000.1). This high-quality genome will serve as a fundamental resource for evolutionary genomics, conservation biology, and breeding of shade-tolerant rice cultivars.

Ribosome biogenesis is critical for plant development and environmental responses. A large number of ribosomal proteins (RPs) and ribosomal biogenesis factors (RBFs) are required for ribosome biogenesis, many of which remain uncharacterized in plants. We report here the identification of Arabidopsis RBF FAN and its interacting partner FAN-INTERACTING PROTEIN 1 (FIP1). As their human and yeast orthologues, FAN-FIP1 interact. Both FAN and FIP1 participate in the processing of pre-rRNAs. Functional loss of FAN or FIP1 knock-down results in developmental retardation and hypersensitivity to heat stresses. We demonstrate that FAN-FIP1 positively mediates brassinosteroid (BR) signaling by ensuring the translation efficiency of the BR receptor-coding gene BRASSINOSTEROID INSENSITIVE 1 (BRI1) through the presence of its upstream open reading frame (uORF). Importantly, BR signaling positively mediates the processing of pre-rRNAs, which may be critical not only for development but also for heat tolerance.

Iron (Fe) is an essential micronutrient for plant growth, and the hormone auxin is a key regulator of developmental processes, including root gravitropism. Here, we investigated the molecular mechanisms underlying the crosstalk between iron nutrition and auxin-mediated root growth in Arabidopsis thaliana. Phenotypic analysis revealed that iron deficiency strongly shaped root system architecture and root gravitropism, and these phenotypes were exacerbated in the iron uptake mutant irt1-1. Genetic analysis revealed that iron deficiency did not aggravate the gravitropic defect of the pin2 mutant, eir1-4, suggesting that iron availability modulates root gravitropism through a PIN2-dependent pathway. Further transcriptomic analysis confirmed that iron deficiency significantly altered the expression of numerous genes related to the auxin pathway, providing molecular evidence for the observed physiological connection. Collectively, this study revealed that iron availability regulates root gravitropic growth by modulating PIN-mediated auxin transport and distribution, providing insights into how plants integrate nutritional cues with developmental programs. Graphical abstract A brief descriptionIron modulates auxin transport and root tip distribution by regulating PIN2 protein, thereby mediating root gravitropism in Arabidopsis. Public summaryO_LIIron nutrition specifically regulates root gravitropism and architecture in Arabidopsis. C_LIO_LIIron deficiency disrupts local auxin homeostasis in root tips and impairs asymmetric distribution in response to gravity. C_LIO_LIIron deficiency stress significantly reduces the abundance of PIN2 protein in root tip cells and disrupts its polar localization pattern on the plasma membrane, thereby precisely modulating polar auxin transport by interfering with the vesicle trafficking and recycling efficiency of PIN2. C_LIO_LIRNA-seq results showed that iron deficiency induced differential expression of multiple auxin-related genes, indicating that iron nutrition affects root development through the auxin pathway. C_LI

The medicinal herb Panax notoginseng produces a structurally diverse array of triterpene saponins (ginsenosides), yet the genetic basis of this chemical complexity remains unclear. Here we present a high-quality chromosome-level genome of diploid P. notoginseng and integrate comparative genomics with multi-tissue, multi-year metabolomics and transcriptomics. Surprisingly, unlike tetraploid Panax species, P. notoginseng shows no general expansion of core saponin biosynthetic gene families. Instead, lineage-specific diversification of UDP-glycosyltransferase (UGT) families, a recent burst of LTR retrotransposons, and enrichment of species-specific genes in metabolic modification pathways point to an alternative evolutionary route. Saponin accumulation follows strict spatiotemporal compartmentalisation, and co-expression network analysis reveals that the biosynthetic machinery is not static but continuously rewired during development-from a basic synthesis module in the first year to a modular pattern supporting both broad accumulation and branch-specific modification by the third year. Seventeen differentially expressed UGTs show clear tissue preferences and saponin-branch correlations. As a representative example, PnUGT33 is tightly linked to the PPD-type saponin branch; structural modelling, molecular docking and 100 ns molecular dynamics simulations demonstrate its differential recognition of diverse triterpene skeletons. Collectively, our findings establish that ginsenoside diversity in diploid P. notoginseng arises primarily from UGT lineage diversification, developmentally rewired regulatory networks and UGT mediated branch selective post-modification, rather than from expansion of core pathway genes. This work provides a new paradigm for understanding how plants achieve metabolic complexity without whole genome duplication or massive gene amplification.

Plants, as sessile organisms, have developed various mechanisms to respond to environmental stress conditions. The plant hormone abscisic acid (ABA) is necessary for the plant to adapt to osmotic stress conditions. However, the molecular mechanisms preceding ABA accumulation remain largely unknown. To isolate transcriptional complexes on the promoter region of NINE-CIS-EPOXYCAROTENOID DIOXYGENASE 3 (NCED3) encoding a rate-limiting enzyme in the ABA biosynthetic pathway in planta, we developed the insertional chromatin immunoprecipitation (iChIP) screen method. The identified ALBA proteins formed condensates through liquid-liquid phase separation (LLPS) in response to osmotic stress conditions. ALBA4 directly binds to stress-inducible genes, including NCED3, and suppresses their stress inducibility. Our results demonstrate how plants respond to osmotic stress at early timepoints before ABA biosynthesis through condensate formation as osmo-sensors.

In tomato plants, the potassium (K) transporter SlHAK5 is integral to root K uptake and overall plant fertility. Under K deficiency, SlHAK5 expression is induced in roots and the encoded transporter is activated via the Ca{superscript 2}-sensing CIPK/CBL complex SlCIPK23/SlCBL1-9. In Arabidopsis, multiple CIPK/CBL complexes can activate AtHAK5, providing alternative regulatory pathways that enhance K uptake. However, the architecture of CIPK/CBL signaling networks has diverged among plant species, necessitating species-specific identification of novel regulatory components. Accordingly, we screened additional tomato CIPK proteins for their capacity to modulate SlHAK5 activity in yeast. SlCIPK15 and SlCIPK26 emerged as potent activators of SlHAK5, acting in concert with SlCBL9. Functional characterization of slcipk15 and slcipk26 mutants revealed that neither contributed significantly to SlHAK5-mediated K uptake in roots. Conversely, both mutants exhibited impaired pollen tube elongation, correlating with reduced K content in pollen relative to wild type. Notably, slcipk26 mutants displayed more severe pollen defects, phenocopying the slhak5 mutant. Further analyses demonstrated that slcipk26 plants suffered compromised seed set and pistil function, paralleling the reproductive deficiencies observed in slhak5 mutants. These findings implicate SlCIPK26 as the principal regulator of SlHAK5 in reproductive tissues. Collectively, our data underscore the role of CIPK paralogs in orchestrating tissue-specific regulation of target proteins, thereby enabling fine-tuned modulation of K transport essential for both vegetative and reproductive development.

Senescence enables plants to remobilize and recycle nutrients from aging organs to support growth, reproduction, and survival. In annual crops like maize, nitrogen remobilization from leaves to grain is incomplete, with 30-50% of nitrogen stranded in aboveground tissues and subject to environmental loss. Mitigating nitrogen loss in annual crops could be achieved by leveraging the physiological strategies of perennial grasses, which remobilize nitrogen and other nutrients into underground organs at the end of the growing season, thereby preventing environmental leakage. To uncover the molecular basis of perennial nitrogen recycling to underground organs, we built a transcriptomic atlas from field-grown plants, comprising 2,685 RNA-seq libraries from 14 grass species within the Panicoideae (Poaceae), utilizing maize and sorghum as annual references for comparative analyses. The atlas spans leaves, roots, stalks, and rhizomes across two seasons, from mid-growing season to senescence. Using a photosynthetic index to align the leafs transition from nitrogen sink to source across species, co-expression network analysis revealed that the subnetworks driving leaf nitrogen recycling are preserved across annuals and perennials. However, we discovered that the subnetworks associated with underground sink establishment, specifically those associated with seed-like dormancy and desiccation tolerance pathways, have diverged among annual crop accessions. Our work identifies conserved gene candidates and networks that could be used to reintroduce perennial-like nutrient recycling into annual crops to enhance long-term nutrient retention in the field.

O_LIResearch on the functions and molecular mechanisms of long non-coding RNAs (lncRNAs) involved in regulating plant freezing tolerance is still in its infancy. Our previous research work identified that lncRNA DROUGHT INDUCED LNCRNA (DRIR) regulates gene expressions in Arabidopsis. However, the underlying molecular mechanism is still unknown. C_LIO_LIThis study demonstrates that lncRNA DRIR regulates plant freezing tolerance by affecting alternative splicing patterns of pre-mRNAs. C_LIO_LIThrough chromatin isolation by RNA purification followed by mass spectrometry (ChIRP-MS), we identified two DRIR interacting proteins: CP29A and CP29B. We showed that the drirD mutant, which exhibits elevated DRIR expression and DRIR overexpression lines showed increased sensitivity to freezing stress, whereas DRIR RNAi lines were more tolerant to the stress. CP29A and CP29B bind to nuclear transcripts and, together with DRIR, regulate pre-mRNA alternative splicing under freezing stress. Notably, DRIR induces the relocalization of CP29A and CP29B to autophagosomes, leading to autophagy-mediated protein degradation. C_LIO_LICollectively, our findings elucidate the molecular mechanism by which DRIR influences the autophagy-based degradation of its binding proteins CP29A and CP29B, thereby regulating plant freezing tolerance by altering the alternative splicing patterns of pre-mRNAs, providing novel insights into the functions and mechanisms of lncRNAs in plants adapting to freezing environments. C_LI

Carotenoid accumulation underlies fruit color and nutritional quality in squash (Cucurbita pepo). One pair of dominant genes, B and L-2, have been long known to interact epistatically, substantially boosting carotenoid accumulation and producing intensely orange-fleshed fruit. However, their molecular identities and regulatory mechanism are unknown. Here, we show that B encodes a truncated H subunit of magnesium chelatase (CpCHLHB) and L-2 encodes a homolog of Arabidopsis Pseudo-Response Regulator 2 (CpAPRR2-A). Significantly, expression of phytoene synthase (CpPSY-A), which encodes the major rate-limiting enzyme in carotenoid biosynthesis, was dramatically upregulated in fruit of B/B L-2/L-2 plants compared with b/b L-2/L-2 or B/B l-2/l-2, showing that the B and L-2 interaction affects CpPSY-A transcription. A similar upregulation was also observed in Arabidopsis gun5 L-2 transgenic plants, where gun5 is a genetic mimic of the C. pepo B gene. The wild-type CpCHLHb physically interacted with CpAPRR2-A, attenuating the CpAPRR2-A-mediated activation of CpPSY-A. In contrast, the truncated CpCHLHB lost its ability to interact with CpAPRR2-A, enabling CpAPRR2-A to activate CpPSY-A and produce intensely orange fruit. These findings uncover the mechanism underlying the epistatic interaction through which B and L-2 act synergistically to boost carotenoid production, offering novel mechanistic insights and key targets for improving crop quality. One-sentence summarySynergistic epistasis between B and L-2 arises from loss of interaction between their encoded proteins, resulting in dramatically upregulating the key rate-limiting enzyme in carotenoid biosynthesis pathway to produce intensely orange-fleshed fruit in squash.

Gibberellins (GAs) influence cell division and elongation, profoundly shaping plant architecture and yield. GA perception occurs when bioactive GAs bind the receptor GID1, promoting DELLA degradation and activating transcriptional programs. While GA signaling in the root endodermis is essential for promoting root elongation, functions of other layers in spatial control of GA responses have not been explored. Here, we developed a synthetic GA (sGA) that does not bind endogenous GID1, together with a modified GID1 (mGID1) engineered to selectively recognize sGA, enabling cell-specific activation of GA signaling in vivo. Using this system in Arabidopsis, we demonstrate that coordinated action of GA signaling in the endodermis, epidermis, and other layers is required for full root elongation. Moreover, cell type-specific expression of GA biosynthetic enzymes indicates the existence of intercellular GA transport. The sGA-mGID1 system provides a versatile platform for spatially precise reprogramming of hormone signaling, enabling synthetic control of developmental processes such as root-shoot growth balance, thereby advancing applications in plant synthetic biology and sustainable crop improvement.

Plants detect neighbours through a reduced red-to-far-red ratio (R:FR), triggering elongation growth that reduces crop yield. Although Gibberellin (GA) is required for the neighbour-proximity (NP) elongation response, bioactive GA levels do not increase sufficiently to account for elongation magnitude, suggesting GA sensitivity as an additional regulated variable. Here, we show that GID1C, one of three Arabidopsis GA receptors, is the primary GA receptor involved in NP-induced elongation. GID1C protein accumulates selectively in hypocotyls and root tips under low R:FR without an increase in bioactive GA. The gid1c mutant shows a reduced elongation response that exogenous GA treatment cannot rescue. Transcriptome profiling reveals that GID1C controls 86% of the NP-responsive transcriptome, including genes for cell growth, division, and transcriptional regulation. Hub analysis identifies ICE1 as a GID1C-repressed transcriptional brake. ICE1 transcript is suppressed under low R:FR in a GID1C-dependent manner, and a phosphorylation-resistant ICE1 allele blocks NP-induced elongation. Together, these findings establish GA perception as an additional regulatory layer in NP, with subfunctionalisation among GID1 paralogs shaping the response to neighbouring plants.