Comparative transcriptome analysis reveals the biocontrol mechanism of Bacillus velezensis E68 against Fusarium graminearum DAOMC 180378, the causal agent of Fusarium head blight.

Fusarium graminearum is the causal agent of Fusarium Head Blight, a serious disease affecting grain crops worldwide. Biological control involves the use of microorganisms to combat plant pathogens such as F. graminearum. Strains of Bacillus velezensis are common biological control candidates for use against F. graminearum and other plant pathogens, as they can secrete antifungal secondary metabolites. Here we study the interaction between B. velezensis E68 and F. graminearum DAOMC 180378 by employing a dual RNA-seq approach to assess the transcriptional changes in both organisms. In dual culture, B. velezensis up-regulated genes related to sporulation and phosphate stress and down-regulated genes related to secondary metabolism, biofilm formation and the tricarboxylic acid cycle. F. graminearum up-regulated genes encoding for killer protein 4-like proteins and genes relating to heavy metal tolerance, and down-regulated genes relating to trichothecene biosynthesis and phenol metabolism. This study provides insight into the molecular mechanisms involved in the interaction between a biocontrol bacterium and a phytopathogenic fungus.

Bacillus velezensis strain B26 modulates the inflorescence and root architecture of Brachypodium distachyon via hormone homeostasis.

Plant growth-promoting rhizobacteria (PGPR) influence plant health. However, the genotypic variations in host organisms affect their response to PGPR. To understand the genotypic effect, we screened four diverse B. distachyon genotypes at varying growth stages for their ability to be colonized by B. velezensis strain B26. We reasoned that B26 may have an impact on the phenological growth stages of B. distachyon genotypes. Phenotypic data suggested the role of B26 in increasing the number of awns and root weight in wild type genotypes and overexpressing transgenic lines. Thus, we characterized the expression patterns of flowering pathway genes in inoculated plants and found that strain B26 modulates the transcript abundance of flowering genes. An increased root volume of inoculated plants was estimated by CT-scanning which suggests the role of B26 in altering the root architecture. B26 also modulated plant hormone homeostasis. A differential response was observed in the transcript abundance of auxin and gibberellins biosynthesis genes in inoculated roots. Our results reveal that B. distachyon plant genotype is an essential determinant of whether a PGPR provides benefit or harm to the host and shed new insight into the involvement of B. velezensis in the expression of flowering genes.

Genotyping by sequencing suggests overwintering of Peronospora destructor in southwestern Québec, Canada.

Several Peronospora species are carried by wind over short and long distances, from warmer climates where they survive on living plants to cooler climates. In eastern Canada, this annual flow of sporangia was thought to be the main source of Peronospora destructor responsible for onion downy mildew. However, the results of a recent study showed that the increasing frequency of onion downy mildew epidemics in eastern Canada is associated with warmer autumns, milder winters, and previous year disease severity, suggesting overwintering of the inoculum in an area where the pathogen is not known to be endogenous. In this study, genotyping by sequencing was used to investigate the population structure of P. destructor at the landscape scale. The study focused on a particular region of southwestern Québec-Les Jardins de Napierville-to determine if the populations were clonal and regionally differentiated. The data were characterized by a high level of linkage disequilibrium, characteristic of clonal organisms. Consequently, the null hypothesis of random mating was rejected when tested on predefined or nonpredefined populations, indicating that linkage disequilibrium was not a function of population structure and suggesting a mixed reproduction mode. Discriminant analysis of principal components performed with predefined population assignment allowed grouping P. destructor isolates by geographical regions, while analysis of molecular variance confirmed that this genetic differentiation was significant at the regional level. Without using a priori population assignment, isolates were clustered into four genetic clusters. These results represent a baseline estimate of the genetic diversity and population structure of P. destructor.

Transcriptional memories mediate the plasticity of cold stress responses to enable morphological acclimation in Brachypodium distachyon.

Plants that successfully acclimate to stress can resume growth under stressful conditions. The grass Brachypodium distachyon can grow a cold-adaptive morphology during cold acclimation. Studies on transcriptional memory (TM) have revealed that plants can be primed for stress by adjusting their transcriptional responses, but the function of TM in stress acclimation is not well understood. We investigated the function of TM during cold acclimation in B. distachyon. Quantitative polymerase chain reaction (qPCR), RNA-seq and chromatin immunoprecipitation qPCR analyses were performed on plants exposed to repeated episodes of cold to characterize the presence and stability of TM during the stress and growth responses of cold acclimation. Transcriptional memory mainly dampened stress responses as growth resumed and as B. distachyon became habituated to cold stress. Although permanent on vernalization gene VRN1, TMs were short-term and reversible on cold-stress genes. Growing under cold conditions also coincided with the acquisition of new and targeted cold-induced transcriptional responses. Overall, TM provided plasticity to cold stress responses during cold acclimation in B. distachyon, leading to stress habituation, acquired stress responses, and resumed growth. Our study shows that chromatin-associated TMs are involved in tuning plant responses to environmental change and, as such, regulate both stress and developmental components that characterize cold-climate adaptation in B. distachyon.

A Crosstalk Between Brachypodium Root Exudates, Organic Acids, and Bacillus velezensis B26, a Growth Promoting Bacterium.

Plant growth-promoting rhizobacteria (PGPR) are associated with plant roots and use organic compounds that are secreted from root exudates as food and energy source. Root exudates can chemoattract and help bacteria to colonize the surface of plant roots by inducing chemotactic responses of rhizospheric bacteria. In this study, we show that root colonization of Brachypodium distachyon by Bacillus velezensis strain B26 depends on several factors. These include root exudates, organic acids, and their biosynthetic genes, chemotaxis, biofilm formation and the induction of biofilm encoding genes. Analysis of root exudates by GC-MS identified five intermediates of the TCA cycle; malic, fumaric, citric, succinic, oxaloacetic acids, and were subsequently evaluated. The strongest chemotactic responses were induced by malic, succinic, citric, and fumaric acids. In comparison, the biofilm formation was induced by all organic acids with maximal induction by citric acid. Relative to the control, the individual organic acids, succinic and citric acids activated the epsD gene related to EPS biofilm, and also the genes encoding membrane protein (yqXM) and hydrophobin component (bslA) of the biofilm of strain B26. Whereas epsA and epsB genes were highly induced genes by succinic acid. Similarly, concentrated exudates released from inoculated roots after 48 h post-inoculation also induced all biofilm-associated genes. The addition of strain B26 to wild type and to icdh mutant line led to a slight induction but not biologically significant relative to their respective controls. Thus, B26 has no effect on the expression of the ICDH gene, both in the wild type and the mutant backgrounds. Our results indicate that root exudates and individual organic acids play an important role in selective recruitment and colonization of PGPR and inducing biofilm. The current study increases the understanding of molecular mechanisms behind biofilm induction by organic acids.

Monitoring of Peronospora destructor Primary and Secondary Inoculum by Real-Time qPCR.

Onion downy mildew (ODM), caused by Peronospora destructor, is a serious threat for onion growers worldwide. In southwestern Québec, Canada, a steady increase in occurrence of ODM has been observed since the mid-2000s. On onion, P. destructor can develop local and systemic infections producing numerous sporangia which act as initial inoculum locally and also for neighboring areas. It also produces oospores capable of surviving in soils and tissues for a prolonged period of time. A recent study showed that ODM epidemics are strongly associated with weather conditions related to production and survival of overwintering inoculum, stressing the need to understand the role of primary (initial) and secondary inoculum. However, P. destructor is an obligate biotrophic pathogen, which complicates the study of inoculum sources. This study aimed at developing a molecular assay specific to P. destructor, allowing its quantification in environmental samples. In this study, a reliable and sensitive hydrolysis probe-based assay multiplexed with an internal control was developed on the internal transcribed spacer (ITS) region to quantify soil- and airborne inoculum of P. destructor. The assay specificity was tested against 17 isolates of P. destructor obtained from different locations worldwide, other members of the order Peronosporales, and various onion pathogens. Validation with artificially inoculated soil and air samples suggested a sensitivity of less than 10 sporangia g-1 of dry soil and 1 sporangium m-3 of air. Validation with environmental air samples shows a linear relationship between microscopic and real-time quantitative PCR counts. In naturally infested soils, inoculum ranged from 0 to 162 sporangia equivalent g-1 of dry soil, which supported the hypothesis of overwintering under northern climates. This assay will be useful for primary and secondary inoculum monitoring to help characterize ODM epidemiology and could be used for daily tactical and short-term strategic decision-making.

Histone tales: lysine methylation, a protagonist in Arabidopsis development.

Histone methylation plays a fundamental role in the epigenetic regulation of gene expression driven by developmental and environmental cues in plants, including Arabidopsis. Histone methyltransferases and demethylases act as 'writers' and 'erasers' of methylation at lysine and/or arginine residues of core histones, respectively. A third group of proteins, the 'readers', recognize and interpret the methylation marks. Emerging evidence confirms the crucial roles of histone methylation in multiple biological processes throughout the plant life cycle. In this review, we summarize the regulatory mechanisms of lysine methylation, especially at histone H3 tails, and focus on the recent advances regarding the roles of lysine methylation in Arabidopsis development, from seed performance to reproductive development, and in callus formation.

Treatment Analogous to Seasonal Change Demonstrates the Integration of Cold Responses in Brachypodium distachyon.

Anthropogenic climate change precipitates the need to understand plant adaptation. Crucial in temperate climates, adaptation to winter is characterized by cold acclimation and vernalization, which respectively lead to freezing tolerance and flowering competence. However, the progression of these responses during fall and their interaction with plant development are not completely understood. By identifying key seasonal cues found in the native range of the cereal model Brachypodium distachyon, we designed a diurnal-freezing treatment (DF) that emulates summer-to-winter change. DF induced unique cold acclimation and vernalization responses characterized by low VERNALIZATION1 (VRN1) expression. Flowering under DF is characterized by an up-regulation of FLOWERING LOCUS T (FT) postvernalization independent of VRN1 expression. DF, while conferring flowering competence, favors a high tolerance to freezing and the development of a winter-hardy plant structure. The findings of this study highlight the contribution of phenotypic plasticity to freezing tolerance and demonstrate the integration of key morphological, physiological, and molecular responses in cold adaptation. The results suggest a fundamental role for VRN1 in regulating cold acclimation, vernalization, and morphological development in B. distachyon This study also establishes the usefulness of reproducing natural cues in laboratory settings.

HvWRKY23 regulates flavonoid glycoside and hydroxycinnamic acid amide biosynthetic genes in barley to combat Fusarium head blight.

Crop plant resistance against pathogens is governed by dynamic molecular and biochemical responses driven by complex transcriptional networks. However, the underlying mechanisms are largely unclear. Here we report an interesting role of HvWRKY23 transcription factor (TF) in modulating defense response against Fusarium head blight (FHB) in barley. The combined approach of gene silencing, metabolomics, real time expression analysis and ab initio bioinformatics tools led to the identification of the HvWRKY23 role in FHB resistance. The knock-down of HvWRKY23 gene in the FHB resistant barley genotype CI9831, followed by inoculation with Fusarium graminearum, led to the down regulation of key flavonoid and hydroxycinnamic acid amide biosynthetic genes resulting in reduced accumulation of resistant related (RR) secondary metabolites such as pelargonidin 3-rutinoside, peonidin 3-rhamnoside-5-glucoside, kaempferol 3-O-arabinoside and other flavonoid glycosides. Reduced abundances of RR metabolites in TF silenced plants were also associated with an increased proportion of spikelets diseased and amount of fungal biomass in spikelets, depicting the role of HvWRKY23 in disease resistance. The luciferase reporter assay demonstrated binding of HvWRKY23 protein to promoters of key flavonoid and hydroxycinnamic acid amides (HCAA) biosynthetic genes, such as HvPAL2, HvCHS1, HvHCT, HvLAC15 and HvUDPGT. The accumulation of high abundances of HCAAs and flavonoid glycosides reinforce cell walls to contain the pathogen to initial infection area. This gene in commercial cultivars can be edited, if non-functional, to enhance resistance against FHB.

Characterization and subcellular localization of histone deacetylases and their roles in response to abiotic stresses in soybean.

BACKGROUND: Histone deacetylases (HDACs) function as key epigenetic factors in repressing the expression of genes in multiple aspects of plant growth, development and plant response to abiotic or biotic stresses. To date, the molecular function of HDACs is well described in Arabidopsis thaliana, but no systematic analysis of this gene family in soybean (Glycine max) has been reported. RESULTS: In this study, 28 HDAC genes from soybean genome were identified, which were asymmetrically distributed on 12 chromosomes. Phylogenetic analysis demonstrated that GmHDACs fall into three major groups previously named RPD3/HDA1, SIR2, and HD2. Subcellular localization analysis revealed that YFP-tagged GmSRT4, GmHDT2 and GmHDT4 were predominantly localized in the nucleus, whereas GmHDA6, GmHDA13, GmHDA14 and GmHDA16 were found in both the cytoplasm and nucleus. Real-time quantitative PCR showed that GmHDA6, GmHDA13, GmHDA14, GmHDA16 and GmHDT4 were broadly expressed across plant tissues, while GmHDA8, GmSRT2, GmSRT4 and GmHDT2 showed differential expression across various tissues. Interestingly, we measured differential changes in GmHDACs transcripts accumulation in response to several abiotic cues, indicating that these epigenetic modifiers could potentially be part of a dynamic transcriptional response to stress in soybean. Finally, we show that the levels of histone marks previously reported to be associated with plant HDACs are modulated by cold and heat in this legume. CONCLUSION: We have identified and classified 28 HDAC genes in soybean. Our data provides insights into the evolution of the HDAC gene family and further support the hypothesis that these genes are important for the plant responses to environmental stress.

Biocontrol Rhizobacterium Pseudomonas sp. 23S Induces Systemic Resistance in Tomato (Solanum lycopersicum L.) Against Bacterial Canker Clavibacter michiganensis subsp. michiganensis.

Tomato bacterial canker disease, caused by Clavibacter michiganensis subsp. michiganensis (Cmm) is a destructive disease and has been a serious concern for tomato industries worldwide. Previously, a rhizosphere isolated strain of Pseudomonas sp. 23S showed antagonistic activity toward Cmm in vitro. This Pseudomonas sp. 23S was characterized to explore the potential of this bacterium for its use in agriculture. Pseudomonas sp. 23S possesses ability to solubilize inorganic phosphorus, and to produce siderophores, indole acetic acid, and hydrogen cyanide. The strain also showed antagonistic activity against Pseudomonas syringae pv. tomato DC 3000. A plant assay indicated that Pseudomonas sp. 23S could promote growth of tomato seedlings. The potential of treating tomato plants with Pseudomonas sp. 23S to reduce the severity of tomato bacterial canker by inducing systemic resistance (ISR) was investigated using well characterized marker genes such as PR1a [salicylic acid (SA)], PI2 [jasmonic acid (JA)], and ACO [ethylene (ET)]. Two-week-old tomato plants were treated with Pseudomonas sp. 23S by soil drench, and Cmm was inoculated into the stem by needle injection on 3, 5, or 7 days post drench. The results indicated that plants treated with Pseudomonas sp. 23S, 5 days prior to Cmm inoculation significantly delayed the progression of the disease. These plants, after 3 weeks from the date of Cmm inoculation, had significantly higher dry shoot and root weight, higher levels of carbon, nitrogen, phosphorus, and potassium in the leaf tissue, and the number of Cmm population in the stem was significantly lower for the plants treated with Pseudomonas sp. 23S. From the real-time quantitative PCR (qRT-PCR) analysis, the treatment with Pseudomonas sp. 23S alone was found to trigger a significant increase in the level of PR1a transcripts in tomato plants. When the plants were treated with Pseudomonas sp. 23S and inoculated with Cmm, the level of PR1a and ACO transcripts were increased, and this response was faster and greater as compared to plants inoculated with Cmm but not treated with Pseudomonas sp. 23S. Overall, the results suggested the involvement of SA signaling pathways for ISR induced by Pseudomonas sp. 23S.

Endophytes of industrial hemp (Cannabis sativa L.) cultivars: identification of culturable bacteria and fungi in leaves, petioles, and seeds.

Plant endophytes are a group of microorganisms that reside asymptomatically within the healthy living tissue. The diversity and molecular and biochemical characterization of industrial hemp-associated endophytes have not been previously studied. This study explored the abundance and diversity of culturable endophytes residing in petioles, leaves, and seeds of three industrial hemp cultivars, and examined their biochemical attributes and antifungal potential. A total of 134 bacterial and 53 fungal strains were isolated from cultivars Anka, CRS-1, and Yvonne. The number of bacterial isolates was similarly distributed among the cultivars, with the majority recovered from petiole tissue. Most fungal strains originated from leaf tissue of cultivar Anka. Molecular and phylogenetic analyses grouped the endophytes into 18 bacterial and 13 fungal taxa, respectively. The most abundant bacterial genera were Pseudomonas, Pantoea, and Bacillus, and the fungal genera were Aureobasidium, Alternaria, and Cochliobolus. The presence of siderophores, cellulase production, and phosphorus solubilization were the main biochemical traits. In proof-of-concept experiments, re-inoculation of tomato roots with some endophytes confirmed their migration to aerial tissues of the plant. Taken together, this study demonstrates that industrial hemp harbours a diversity of microbial endophytes, some of which could be used in growth promotion and (or) in biological control designed experiments.

Diversification of the Histone Acetyltransferase GCN5 through Alternative Splicing in Brachypodium distachyon.

The epigenetic modulatory SAGA complex is involved in various developmental and stress responsive pathways in plants. Alternative transcripts of the SAGA complex's enzymatic subunit GCN5 have been identified in Brachypodium distachyon. These splice variants differ based on the presence and integrity of their conserved domain sequences: the histone acetyltransferase domain, responsible for catalytic activity, and the bromodomain, involved in acetyl-lysine binding and genomic loci targeting. GCN5 is the wild-type transcript, while alternative splice sites result in the following transcriptional variants: L-GCN5, which is missing the bromodomain and S-GCN5, which lacks the bromodomain as well as certain motifs of the histone acetyltransferase domain. Absolute mRNA quantification revealed that, across eight B. distachyon accessions, GCN5 was the dominant transcript isoform, accounting for up to 90% of the entire transcript pool, followed by L-GCN5 and S-GCN5. A cycloheximide treatment further revealed that the S-GCN5 splice variant was degraded through the nonsense-mediated decay pathway. All alternative BdGCN5 transcripts displayed similar transcript profiles, being induced during early exposure to heat and displaying higher levels of accumulation in the crown, compared to aerial tissues. All predicted protein isoforms localize to the nucleus, which lends weight to their purported epigenetic functions. S-GCN5 was incapable of forming an in vivo protein interaction with ADA2, the transcriptional adaptor that links the histone acetyltransferase subunit to the SAGA complex, while both GCN5 and L-GCN5 interacted with ADA2, which suggests that a complete histone acetyltransferase domain is required for BdGCN5-BdADA2 interaction in vivo. Thus, there has been a diversification in BdGCN5 through alternative splicing that has resulted in differences in conserved domain composition, transcript fate and in vivo protein interaction partners. Furthermore, our results suggest that B. distachyon may harbor compositionally distinct SAGA-like complexes that differ based on their histone acetyltransferase subunit.

Accelerated Growth Rate and Increased Drought Stress Resilience of the Model Grass Brachypodium distachyon Colonized by Bacillus subtilis B26.

Plant growth-promoting bacteria (PGB) induce positive effects in plants, for instance, increased growth and reduced abiotic stresses susceptibility. The mechanisms by which these bacteria impact the host plant are numerous, diverse and often specific. Here, we studied the agronomical, molecular and biochemical effects of the endophytic PGB Bacillus subtilis B26 on the full life cycle of Brachypodium distachyon Bd21, an established model species for functional genomics in cereal crops and temperate grasses. Inoculation of Brachypodium with B. subtilis strain B26 increased root and shoot weights, accelerated growth rate and seed yield as compared to control plants. B. subtilis strain B26 efficiently colonized the plant and was recovered from roots, stems and blades as well as seeds of Brachypodium, indicating that the bacterium is able to migrate, spread systemically inside the plant, establish itself in the aerial plant tissues and organs, and is vertically transmitted to seeds. The presence of B. subtilis strain B26 in the seed led to systemic colonization of the next generation of Brachypodium plants. Inoculated Brachypodium seedlings and mature plants exposed to acute and chronic drought stress minimized the phenotypic effect of drought compared to plants not harbouring the bacterium. Protection from the inhibitory effects of drought by the bacterium was linked to upregulation of the drought-response genes, DREB2B-like, DHN3-like and LEA-14-A-like and modulation of the DNA methylation genes, MET1B-like, CMT3-like and DRM2-like, that regulate the process. Additionally, total soluble sugars and starch contents increased in stressed inoculated plants, a biochemical indication of drought tolerance. In conclusion, we show a single inoculation of Brachypodium with a PGB affected the whole growth cycle of the plant, accelerating its growth rates, shortening its vegetative period, and alleviating drought stress effects. These effects are relevant to grasses and cereal crops.

Cold acclimation induces distinctive changes in the chromatin state and transcript levels of COR genes in Cannabis sativa varieties with contrasting cold acclimation capacities.

Little is known about the capacity of Cannabis sativa to cold-acclimate and develop freezing tolerance. This study investigates the cold acclimation (CA) capacity of nine C. sativa varieties and the underlying genetic and epigenetic responses. The varieties were divided into three groups based on their contrasting CA capacities by comparing the survival of non-acclimated and cold-acclimated plants in whole-plant freeze tests. In response to the CA treatment, all varieties accumulated soluble sugars but only the varieties with superior capacity for CA could maintain higher levels throughout the treatment. In addition, the varieties that acclimated most efficiently accumulated higher transcript levels of cold-regulated (COR) genes and genes involved in de novo DNA methylation while displaying locus- and variety-specific changes in the levels of H3K9ac, H3K27me3 and methylcytosine (MeC) during CA. Furthermore, these hardy C. sativa varieties displayed significant increases in MeC levels at COR gene loci when deacclimated, suggesting a role for locus-specific DNA methylation in deacclimation. This study uncovers the molecular mechanisms underlying CA in C. sativa and reveals higher levels of complexity regarding how genetic, epigenetic and environmental factors intertwine.

Photoreceptor partner FHY1 has an independent role in gene modulation and plant development under far-red light.

To incorporate the far-red light (FR) signal into a strategy for optimizing plant growth, FAR-RED ELONGATED HYPOCOTYL1 (FHY1) mediates the nuclear translocation of the FR photoreceptor phytochrome A (phyA) and facilitates the association of phyA with the promoters of numerous associated genes crucial for the response to environmental stimuli. However, whether FHY1 plays additional roles after FR irradiation remains elusive. Here, through the global identification of FHY1 chromatin association sites through ChIP-seq analysis and by the comparison of FHY1-associated sites with phyA-associated sites, we demonstrated that nuclear FHY1 can either act independently of phyA or act in association with phyA to activate the expression of distinct target genes. We also determined that phyA can act independently of FHY1 in regulating phyA-specific target genes. Furthermore, we determined that the independent FHY1 nuclear pathway is involved in crucial aspects of plant development, as in the case of inhibited seed germination under FR during salt stress. Notably, the differential presence of cis-elements and transcription factors in common and unique FHY1- and/or phyA-associated genes are indicative of the complexity of the independent and coordinated FHY1 and phyA pathways. Our study uncovers previously unidentified aspects of FHY1 function beyond its currently recognized role in phyA-dependent photomorphogenesis.

Arabidopsis Phytochrome A Directly Targets Numerous Promoters for Individualized Modulation of Genes in a Wide Range of Pathways.

The far-red light (FR) photoreceptor phytochrome A (phyA) contains no DNA binding domain but associates with the CHALCONE SYNTHASE promoter through its chaperone FAR-RED ELONGATED HYPOCOTYL1 and transcription factors. Here, we performed a genome-wide identification of phyA targets using a combination of phyA chromatin immunoprecipitation and RNA sequencing methods in Arabidopsis thaliana. Our results indicate that phyA signaling widely affects gene promoters involved in multiple FR-modulated aspects of plant growth. Furthermore, we observed an enrichment of hormone- and stress-responsive elements in the phyA direct target promoters, indicating that a much broader than expected range of transcription factors is involved in the phyA signaling pathway. To verify our hypothesis that phyA regulates genes other than light-responsive ones through the interaction with corresponding transcription factors, we examined the action of phyA on one of its direct target genes, NAC019, which encodes an abscisic acid-dependent transcription factor. The phyA signaling cascade not only targets two G-boxes on the NAC019 promoter for subsequent transcriptional regulation but also positively coordinates with the abscisic acid signaling response for root elongation inhibition under FR. Our study provides new insight into how plants rapidly fine-tune their growth strategy upon changes in the light environment by escorting photoreceptors to the promoters of hormone- or stress-responsive genes for individualized modulation.

Comparative analysis of the cold acclimation and freezing tolerance capacities of seven diploid Brachypodium distachyon accessions.

BACKGROUND AND AIMS: Cold is a major constraint for cereal cultivation under temperate climates. Winter-hardy plants interpret seasonal changes and can acquire the ability to resist sub-zero temperatures. This cold acclimation process is associated with physiological, biochemical and molecular alterations in cereals. Brachypodium distachyon is considered a powerful model system to study the response of temperate cereals to adverse environmental conditions. To date, little is known about the cold acclimation and freezing tolerance capacities of Brachypodium. The main objective of this study was to evaluate the cold hardiness of seven diploid Brachypodium accessions. METHODS: An integrated approach, involving monitoring of phenological indicators along with expression profiling of the major vernalization regulator VRN1 orthologue, was followed. In parallel, soluble sugars and proline contents were determined along with expression profiles of two COR genes in plants exposed to low temperatures. Finally, whole-plant freezing tests were performed to evaluate the freezing tolerance capacity of Brachypodium. KEY RESULTS: Cold treatment accelerated the transition from the vegetative to the reproductive phase in all diploid Brachypodium accessions tested. In addition, low temperature exposure triggered the gradual accumulation of BradiVRN1 transcripts in all accessions tested. These accessions exhibited a clear cold acclimation response by progressively accumulating proline, sugars and COR gene transcripts. However, whole-plant freezing tests revealed that these seven diploid accessions only have a limited capacity to develop freezing tolerance when compared with winter varieties of temperate cereals such as wheat and barley. Furthermore, little difference in terms of survival was observed among the accessions tested despite their previous classification as either spring or winter genotypes. CONCLUSIONS: This study is the first to characterize the freezing tolerance capacities of B. distachyon and provides strong evidence that some diploid accessions such as Bd21 have a facultative growth habit.