Regulatory networks in plant responses to drought and cold stress.

Drought and cold represent distinct types of abiotic stress, each initiating unique primary signaling pathways in response to dehydration and temperature changes, respectively. However, a convergence at the gene regulatory level is observed where a common set of stress-responsive genes is activated to mitigate the impacts of both stresses. In this review, we explore these intricate regulatory networks, illustrating how plants coordinate distinct stress signals into a collective transcriptional strategy. We delve into the molecular mechanisms of stress perception, stress signaling, and the activation of gene regulatory pathways, with a focus on insights gained from model species. By elucidating both the shared and distinct aspects of plant responses to drought and cold, we provide insight into the adaptive strategies of plants, paving the way for the engineering of stress-resilient crop varieties that can withstand a changing climate.

Genetic variation for grain nutritional profile and yield potential in sorghum and the possibility of selection for drought tolerance under irrigated conditions.

BACKGROUND: Increasing grain nutritional value in sorghum (Sorghum bicolor) is a paramount breeding objective, as is increasing drought resistance (DR), because sorghum is grown mainly in drought-prone areas. The genetic basis of grain nutritional traits remains largely unknown. Marker-assisted selection using significant loci identified through genome-wide association study (GWAS) shows potential for selecting desirable traits in crops. This study assessed natural variation available in sorghum accessions from around the globe to identify novel genes or genomic regions with potential for improving grain nutritional value, and to study associations between DR traits and grain weight and nutritional composition. RESULTS: We dissected the genetic architecture of grain nutritional composition, protein content, thousand-kernel weight (TKW), and plant height (PH) in sorghum through GWAS of 163 unique African and Asian accessions under irrigated and post-flowering drought conditions. Several QTLs were detected. Some were significantly associated with DR, TKW, PH, protein, and Zn, Mn, and Ca contents. Genomic regions on chromosomes 1, 2, 4, 8, 9, and 10 were associated with TKW, nutritional, and DR traits; colocalization patterns of these markers indicate potential for simultaneous improvement of these traits. In African accessions, markers associated with TKW were mapped to six regions also associated with protein, Zn, Ca, Mn, Na, and DR, suggesting the potential for simultaneous selection for higher grain nutrition and TKW. Our results indicate that it may be possible to select for increased DR on the basis of grain nutrition and weight potential. CONCLUSIONS: This study provides a valuable resource for selecting landraces for use in plant breeding programs and for identifying loci that may contribute to grain nutrition and weight with the hope of producing cultivars that combine improved yield traits, nutrition, and DR.

Metabolic and transcriptomic profiling during wheat seed development under progressive drought conditions.

Globally, bread wheat (Triticum aestivum) is one of the most important staple foods; when exposed to drought, wheat yields decline. Although much research has been performed to generate higher yield wheat cultivars, there have been few studies on improving end-product quality under drought stress, even though wheat is processed into flour to produce so many foods, such as bread, noodles, pancakes, cakes, and cookies. Recently, wheat cultivation has been affected by severe drought caused by global climate change. In previous studies, seed shrinkage was observed in wheat exposed to continuous drought stress during seed development. In this study, we investigated how progressive drought stress affected seed development by metabolomic and transcriptomic analyses. Metabolite profiling revealed the drought-sensitive line reduced accumulation of proline and sugar compared with the water-saving, drought-tolerant transgenic line overexpressing the abscisic acid receptor TaPYL4 under drought conditions in spikelets with developing seeds. Meanwhile, the expressions of genes involved in translation, starch biosynthesis, and proline and arginine biosynthesis was downregulated in the drought-sensitive line. These findings suggest that seed shrinkage, exemplifying a deficiency in endosperm, arose from the hindered biosynthesis of crucial components including seed storage proteins, starch, amino acids, and sugars, ultimately leading to their inadequate accumulation within spikelets. Water-saving drought tolerant traits of wheat would aid in supporting seed formation under drought conditions.

A metabolome genome-wide association study implicates histidine N-pi-methyltransferase as a key enzyme in N-methylhistidine biosynthesis in Arabidopsis thaliana.

A genome-wide association study (GWAS), which uses information on single nucleotide polymorphisms (SNPs) from many accessions, has become a powerful approach to gene identification. A metabolome GWAS (mGWAS), which relies on phenotypic information based on metabolite accumulation, can identify genes that contribute to primary and secondary metabolite contents. In this study, we carried out a mGWAS using seed metabolomic data from Arabidopsis thaliana accessions obtained by liquid chromatography-mass spectrometry to identify SNPs highly associated with the contents of metabolites such as glucosinolates. These SNPs were present in genes known to be involved in glucosinolate biosynthesis, thus confirming the effectiveness of our analysis. We subsequently focused on SNPs detected in an unknown methyltransferase gene associated with N-methylhistidine content. Knockout and overexpression of A. thaliana lines of this gene had significantly decreased and increased N-methylhistidine contents, respectively. We confirmed that the overexpressing line exclusively accumulated histidine methylated at the pi position, not at the tau position. Our findings suggest that the identified methyltransferase gene encodes a key enzyme for N-methylhistidine biosynthesis in A. thaliana.

Exome-wide variation in a diverse barley panel reveals genetic associations with ten agronomic traits in Eastern landraces.

Barley (Hordeum vulgare ssp. vulgare) is one of the first crops to be domesticated and is adapted to a wide range of environments. Worldwide barley germplasm collections possess valuable allelic variations that could further improve barley productivity. Although barley genomics has offered a global picture of allelic variation among varieties and its association with various agronomic traits, polymorphisms from East Asian varieties remain scarce. In this study, we analyze exome polymorphisms in a panel of 274 barley varieties collected worldwide, including 137 varieties from East Asian countries and Ethiopia. We reveal the underlying population structure and conduct genome-wide association studies for 10 agronomic traits. Moreover, we examin genome-wide associations for traits related to grain size such as awn length and glume length. Our results demonstrate the value of diverse barley germplasm panels containing Eastern varieties, highlighting their distinct genomic signatures relative to Western subpopulations.

ER Stress and the Unfolded Protein Response: Homeostatic Regulation Coordinate Plant Survival and Growth.

The endoplasmic reticulum (ER), a eukaryotic organelle, is the major site of protein biosynthesis. The disturbance of ER function by biotic or abiotic stress triggers the accumulation of misfolded or unfolded proteins in the ER. The unfolded protein response (UPR) is the best-studied ER stress response. This transcriptional regulatory system senses ER stress, activates downstream genes that function to mitigate stress, and restores homeostasis. In addition to its conventional role in stress responses, recent reports indicate that the UPR is involved in plant growth and development. In this review, we summarize the current knowledge of ER stress sensing and the activation and downstream regulation of the UPR. We also describe how the UPR modulates both plant growth and stress tolerance by maintaining ER homeostasis. Lastly, we propose that the UPR is a major component of the machinery that balances the trade-off between plant growth and survival in a dynamic environment.

Time-series transcriptome of Brachypodium distachyon during bacterial flagellin-induced pattern-triggered immunity.

Plants protect themselves from microorganisms by inducing pattern-triggered immunity (PTI) via recognizing microbe-associated molecular patterns (MAMPs), conserved across many microbes. Although the MAMP perception mechanism and initial events during PTI have been well-characterized, knowledge of the transcriptomic changes in plants, especially monocots, is limited during the intermediate and terminal stages of PTI. Here, we report a time-series high-resolution RNA-sequencing (RNA-seq) analysis during PTI in the leaf disks of Brachypodium distachyon. We identified 6,039 differentially expressed genes (DEGs) in leaves sampled at 0, 0.5, 1, 3, 6, and 12 hours after treatment (hat) with the bacterial flagellin peptide flg22. The k-means clustering method classified these DEGs into 10 clusters (6 upregulated and 4 downregulated). Based on the results, we selected 10 PTI marker genes in B. distachyon. Gene ontology (GO) analysis suggested a tradeoff between defense responses and photosynthesis during PTI. The data indicated the recovery of photosynthesis started at least at 12 hat. Over-representation analysis of transcription factor genes and cis-regulatory elements in DEG promoters implied the contribution of 12 WRKY transcription factors in plant defense at the early stage of PTI induction.

Admixture of divergent genomes facilitates hybridization across species in the family Brassicaceae.

Hybridization and polyploidization are pivotal to plant evolution. Genetic crosses between distantly related species are rare in nature due to reproductive barriers but how such hurdles can be overcome is largely unknown. Here we report the hybrid genome structure of xBrassicoraphanus, a synthetic allotetraploid of Brassica rapa and Raphanus sativus. We performed cytogenetic analysis and de novo genome assembly to examine chromosome behaviors and genome integrity in the hybrid. Transcriptome analysis was conducted to investigate expression of duplicated genes in conjunction with epigenome analysis to address whether genome admixture entails epigenetic reconfiguration. Allotetraploid xBrassicoraphanus retains both parental chromosomes without genome rearrangement. Meiotic synapsis formation and chromosome exchange are avoided between nonhomologous progenitor chromosomes. Reconfiguration of transcription network occurs, and less divergent cis-elements of duplicated genes are associated with convergent expression. Genome-wide DNA methylation asymmetry between progenitors is largely maintained but, notably, B. rapa-originated transposable elements are transcriptionally silenced in xBrassicoraphanus through gain of DNA methylation. Our results demonstrate that hybrid genome stabilization and transcription compatibility necessitate epigenome landscape adjustment and rewiring of cis-trans interactions. Overall, this study suggests that a certain extent of genome divergence facilitates hybridization across species, which may explain the great diversification and expansion of angiosperms during evolution.

Arabidopsis TBP-ASSOCIATED FACTOR 12 ortholog NOBIRO6 controls root elongation with unfolded protein response cofactor activity.

Plant root growth is indeterminate but continuously responds to environmental changes. We previously reported on the severe root growth defect of a double mutant in bZIP17 and bZIP28 (bz1728) modulating the unfolded protein response (UPR). To elucidate the mechanism by which bz1728 seedlings develop a short root, we obtained a series of bz1728 suppressor mutants, called nobiro, for rescued root growth. We focused here on nobiro6, which is defective in the general transcription factor component TBP-ASSOCIATED FACTOR 12b (TAF12b). The expression of hundreds of genes, including the bZIP60-UPR regulon, was induced in the bz1728 mutant, but these inductions were markedly attenuated in the bz1728nobiro6 mutant. In view of this, we assigned transcriptional cofactor activity via physical interaction with bZIP60 to NOBIRO6/TAF12b. The single nobiro6/taf12b mutant also showed an altered sensitivity to endoplasmic reticulum stress for both UPR and root growth responses, demonstrating that NOBIRO6/TAF12b contributes to environment-responsive root growth control through UPR.

DNA demethylase ROS1 prevents inheritable DREB1A/CBF3 repression by transgene-induced promoter methylation in the Arabidopsis ice1-1 mutant.

KEY MESSAGE: In the ros1-defective mutant, DREB1A repression by the transgene-induced promoter methylation of ice1-1 became inheritable across generations even in the absence of the causative transgene NICE1. Transgene silencing (TGS) is a widely observed event during plant bioengineering, which is presented as a gradual decrease in ectopic gene expression across generations and occasionally coupled with endogenous gene silencing based on DNA sequence similarity. TGS is known to be established by guided DNA methylation machinery. However, the machinery underlying gene recovery from TGS has not been fully elucidated. We previously reported that in ice1-1 outcross descendants, the expressional repression and recovery of DREB1A/CBF3 were instantly achieved by a newly discovered NICE1 transgene, instead of the formerly proposed ice1-1 mutation in the ICE1 gene. The plants harboring NICE1 produced small RNAs targeting and causing the DREB1A promoter to be hypermethylated and silenced. To analyze the role of the plant-specific active DNA demethylase REPRESSOR OF SILENCING 1 (ROS1) in instant DREB1A recovery, we propagated the NICE1-segregating population upon ros1 dysfunction and evaluated the gene expression and DNA methylation levels of DREB1A through generations. Our results showed that the epigenetic DREB1A repression was substantially sustained in subsequent generations even without NICE1 and stably inherited across generations. Consistent with the gene expression results, only incomplete DNA methylation removal was detected in the same generations. These results indicate that a novel inheritable epiallele emerged by the ros1 dysfunction. Overall, our study reveals the important role of ROS1 in the inheritability of TGS-associated gene repression.

DREB1A/CBF3 Is Repressed by Transgene-Induced DNA Methylation in the Arabidopsis ice1 -1 Mutant.

DREB1/CBFs are key transcription factors involved in plant cold stress adaptation. The expression of DREB1/CBFs triggers a cold-responsive transcriptional cascade, after which many stress tolerance genes are expressed. Thus, elucidating the mechanisms of cold stress-inducible DREB1/CBF expression is important to understand the molecular mechanisms of plant cold stress responses and tolerance. We analyzed the roles of a transcription factor, INDUCER OF CBF EXPRESSION1 (ICE1), that is well known as an important transcriptional activator in the cold-inducible expression of DREB1A/CBF3 in Arabidopsis (Arabidopsis thaliana). ice1-1 is a widely accepted mutant allele known to abolish cold-inducible DREB1A expression, and this evidence has strongly supported ICE1-DREB1A regulation for many years. However, in ice1-1 outcross descendants, we unexpectedly discovered that ice1-1 DREB1A repression was genetically independent of the ice1-1 allele ICE1(R236H). Moreover, neither ICE1 overexpression nor double loss-of-function mutation of ICE1 and its homolog SCRM2 altered DREB1A expression. Instead, a transgene locus harboring a reporter gene in the ice1-1 genome was responsible for altering DREB1A expression. The DREB1A promoter was hypermethylated due to the transgene. We showed that DREB1A repression in ice1-1 results from transgene-induced silencing and not genetic regulation by ICE1. The ICE1(R236H) mutation has also been reported as scrm-D, which confers constitutive stomatal differentiation. The scrm-D phenotype and the expression of a stomatal differentiation marker gene were confirmed to be linked to the ICE1(R236H) mutation. We propose that the current ICE1-DREB1 regulatory model should be revalidated without the previous assumptions.

Tuning water-use efficiency and drought tolerance in wheat using abscisic acid receptors.

Water availability is a key determinant of terrestrial plant productivity. Many climate models predict that water stress will increasingly challenge agricultural yields and exacerbate projected food deficits. To ensure food security and increase agricultural efficiency, crop water productivity must be increased. Research over past decades has established that the phytohormone abscisic acid (ABA) is a central regulator of water use and directly regulates stomatal opening and transpiration. In this study, we investigated whether the water productivity of wheat could be improved by increasing its ABA sensitivity. We show that overexpression of a wheat ABA receptor increases wheat ABA sensitivity, which significantly lowers a plant's lifetime water consumption. Physiological analyses demonstrated that this water-saving trait is a consequence of reduced transpiration and a concomitant increase in photosynthetic activity, which together boost grain production per litre of water and protect productivity during water deficit. Our findings provide a general strategy for increasing water productivity that should be applicable to other crops because of the high conservation of the ABA signalling pathway.

ROS1-Dependent DNA Demethylation Is Required for ABA-Inducible NIC3 Expression.

DNA methylation plays an important role in diverse developmental processes in many eukaryotes, including the response to environmental stress. Abscisic acid (ABA) is a plant hormone that is up-regulated under stress. The involvement of DNA methylation in the ABA response has been reported but is poorly understood. DNA demethylation is a reverse process of DNA methylation and often induces structural changes of chromatin leading to transcriptional activation. In Arabidopsis (Arabidopsis thaliana), active DNA demethylation depends on the activity of REPRESSOR OF SILENCING 1 (ROS1), which directly excises 5-methylcytosine from DNA. Here we showed that ros1 mutants were hypersensitive to ABA during early seedling development and root elongation. Expression levels of some ABA-inducible genes were decreased in ros1 mutants, and more than 60% of their proximal regions became hypermethylated, indicating that a subset of ABA-inducible genes are under the regulation of ROS1-dependent DNA demethylation. Notable among them is NICOTINAMIDASE 3 (NIC3) that encodes an enzyme that converts nicotinamide to nicotinic acid in the NAD+ salvage pathway. Many enzymes in this pathway are known to be involved in stress responses. The nic3 mutants display hypersensitivity to ABA, whereas overexpression of NIC3 restores normal ABA responses. Our data suggest that NIC3 is responsive to ABA but requires ROS1-mediated DNA demethylation at the promoter as a prerequisite to transcriptional activation. These findings suggest that ROS1-induced active DNA demethylation maintains the active state of NIC3 transcription in response to ABA.

MYB1 transcription factor is a candidate responsible for red root skin in radish (Raphanus sativus L.).

Root skin color is one of the economically important traits in radish (Raphanus sativus), and the pigmentation in red skin varieties is largely attributable to anthocyanin accumulation. Pelargonidin was found as a major anthocyanin pigment accumulated in the sub-epidermal layer of red radish roots. In the 20 F2 population generated from the F1 with red root skins, root skins with red and white colors segregated in a 3:1 ratio. Additionally, a test cross between a red F3 individual and a white skin individual gave rise to 1:1 segregation of red and white, indicating that the root skin color of radish is determined by a single locus and red color is dominant over white. We performed association mapping for root skin color using SNPs obtained from RNA-seq analysis. Segregation analysis on the 152 F3 test-cross population revealed an RsMyb1 transcription factor as a candidate gene to determine root skin color. A PCR marker based on the polymorphism within 2 kb of RsMyb1 was developed and tested on 12 and 152 individuals from F2 and F3 test cross populations, respectively, and red and white root skin colors were completely distinguished corresponding to the genotypes. Expression levels of RsMyb1 in red or purple root cultivars were significantly higher than in white root cultivars. These findings suggest that RsMyb1 is a crucial determinant for anthocyanin biosynthesis in radish roots, and the molecular marker developed in this study will be useful for marker-assisted selection for red skin individuals at early seedling stages.

A population of wheat multiple synthetic derivatives: an effective platform to explore, harness and utilize genetic diversity of Aegilops tauschii for wheat improvement.

KEY MESSAGE: The multiple synthetic derivatives platform described in this study will provide an opportunity for effective utilization of Aegilops tauschii traits and genes for wheat breeding. Introducing genes from wild relatives is the best option to increase genetic diversity and discover new alleles necessary for wheat improvement. A population harboring genomic fragments from the diploid wheat progenitor Aegilops tauschii Coss. in the background of bread wheat (Triticum aestivum L.) was developed by crossing and backcrossing 43 synthetic wheat lines with the common wheat cultivar Norin 61. We named this population multiple synthetic derivatives (MSD). To validate the suitability of this population for wheat breeding and genetic studies, we randomly selected 400 MSD lines and genotyped them by using Diversity Array Technology sequencing markers. We scored black glume as a qualitative trait and heading time in two environments in Sudan as a quantitative trait. Our results showed high genetic diversity and less recombination which is expected from the nature of the population. Genome-wide association (GWA) analysis showed one QTL at the short arm of chromosome 1D different from those alleles reported previously indicating that black glume in the MSD population is controlled by new allele at the same locus. For heading time, from the two environments, GWA analysis revealed three QTLs on the short arms of chromosomes 2A, 2B and 2D and two on the long arms of chromosomes 5A and 5D. Using the MSD population, which represents the diversity of 43 Ae. tauschii accessions representing most of its natural habitat, QTLs or genes and desired phenotypes (such as drought, heat and salinity tolerance) could be identified and selected for utilization in wheat breeding.

Efficient anchoring of alien chromosome segments introgressed into bread wheat by new Leymus racemosus genome-based markers.

BACKGROUND: The tertiary gene pool of bread wheat, to which Leymus racemosus belongs, has remained underutilized due to the current limited genomic resources of the species that constitute it. Continuous enrichment of public databases with useful information regarding these species is, therefore, needed to provide insights on their genome structures and aid successful utilization of their genes to develop improved wheat cultivars for effective management of environmental stresses. RESULTS: We generated de novo DNA and mRNA sequence information of L. racemosus and developed 110 polymorphic PCR-based markers from the data, and to complement the PCR markers, DArT-seq genotyping was applied to develop additional 9990 SNP markers. Approximately 52% of all the markers enabled us to clearly genotype 22 wheat-L. racemosus chromosome introgression lines, and L. racemosus chromosome-specific markers were highly efficient in detailed characterization of the translocation and recombination lines analyzed. A further analysis revealed remarkable transferability of the PCR markers to three other important Triticeae perennial species: L. mollis, Psathyrostachys huashanica and Elymus ciliaris, indicating their suitability for characterizing wheat-alien chromosome introgressions carrying chromosomes of these genomes. CONCLUSION: The efficiency of the markers in characterizing wheat-L. racemosus chromosome introgression lines proves their reliability, and their high transferability further broadens their scope of application. This is the first report on sequencing and development of markers from L. racemosus genome and the application of DArT-seq to develop markers from a perennial wild relative of wheat, marking a paradigm shift from the seeming concentration of the technology on cultivated species. Integration of these markers with appropriate cytogenetic methods would accelerate development and characterization of wheat-alien chromosome introgression lines.

ER-Anchored Transcription Factors bZIP17 and bZIP28 Regulate Root Elongation.

The unfolded protein response (UPR) is a eukaryotic transcriptional regulatory network that is activated upon the accumulation of malformed proteins in the endoplasmic reticulum (ER). In Arabidopsis (Arabidopsis thaliana), three bZIP transcription factors modulate the UPR: bZIP17, bZIP28, and bZIP60. Although bZIP28 and bZIP60 have been relatively well studied, the physiological and transcriptional roles of bZIP17 remain largely unknown. Here, we generated a double knockout mutant of bZIP17 and bZIP28 to elucidate the function of bZIP17. The mutant plant exhibited multiple developmental defects, including markedly reduced root elongation and constantly overinduced bZIP60 activity, indicating the essential roles of bZIP17 and bZIP28 in plant development and UPR modulation. Extended analysis of the transcriptomes of three double knockout mutants of bZIP17, bZIP28, and bZIP60 revealed that bZIP28 and bZIP60 are the major activators of the canonical induced UPR. By contrast, bZIP17 functions with bZIP28 to mediate the noninducible expression of multiple genes involved in cell growth, particularly to sustain their expression under stress conditions. Our study reveals pivotal roles of bZIP17 in the plant UPR and vegetative development, with functional redundancy to bZIP28.

BPM-CUL3 E3 ligase modulates thermotolerance by facilitating negative regulatory domain-mediated degradation of DREB2A in Arabidopsis.

DEHYDRATION-RESPONSIVE ELEMENT BINDING PROTEIN 2A (DREB2A) acts as a key transcription factor in both drought and heat stress tolerance in Arabidopsis and induces the expression of many drought- and heat stress-inducible genes. Although DREB2A expression itself is induced by stress, the posttranslational regulation of DREB2A, including protein stabilization, is required for its transcriptional activity. The deletion of a 30-aa central region of DREB2A known as the negative regulatory domain (NRD) transforms DREB2A into a stable and constitutively active form referred to as DREB2A CA. However, the molecular basis of this stabilization and activation has remained unknown for a decade. Here we identified BTB/POZ AND MATH DOMAIN proteins (BPMs), substrate adaptors of the Cullin3 (CUL3)-based E3 ligase, as DREB2A-interacting proteins. We observed that DREB2A and BPMs interact in the nuclei, and that the NRD of DREB2A is sufficient for its interaction with BPMs. BPM-knockdown plants exhibited increased DREB2A accumulation and induction of DREB2A target genes under heat and drought stress conditions. Genetic analysis indicated that the depletion of BPM expression conferred enhanced thermotolerance via DREB2A stabilization. Thus, the BPM-CUL3 E3 ligase is likely the long-sought factor responsible for NRD-dependent DREB2A degradation. Through the negative regulation of DREB2A stability, BPMs modulate the heat stress response and prevent an adverse effect of excess DREB2A on plant growth. Furthermore, we found the BPM recognition motif in various transcription factors, implying a general contribution of BPM-mediated proteolysis to divergent cellular responses via an accelerated turnover of transcription factors.

RNA-Seq using bulked recombinant inbred line populations uncovers the importance of brassinosteroid for seed longevity after priming treatments.

Seed priming is a commercially used technique for improving seed performance including germination. However, the treatment sometimes reduces seed longevity as a side effect, limiting the storable period or longevity of the seeds. To overcome this problem, molecular mechanisms involved in the loss of seed longevity during priming were analyzed using natural variations of Arabidopsis thaliana. We found that the Est-1 accession retained longevity for longer after priming compared to the reference accession Col-0. QTL analysis using 279 recombinant inbred lines (RILs) derived from the Est-1 × Col-0 detected three QTL regions associated with the loss of seed longevity during priming. Bulked transcriptome analysis (RNA-Seq with bulked RIL populations) revealed that genes related to brassinosteroid (BR) biosynthesis/signaling and cell wall modification were highly expressed in primed seeds with shorter longevity. After priming, BR-deficient mutants cyp85a1/a2 and det2 showed significantly longer longevity than the wild type (WT). Moreover, tetrazolium staining indicated that mutant seed coats were less permeable after priming than those of WT. We suggest that the loss of seed longevity in primed seed is due to increased seed coat permeability, which is positively regulated, at least partly, via BR signaling.