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Measurement of the surface free energy (SFE) of a material allows the prediction of its adhesion properties.

International audience; Interactions among plants have been long recognized as a major force driving plant community dynamics and crop yield. Surprisingly, our knowledge of the ecological genetics associated with variation of plant-plant interactions remains limited. In this opinion article by scientists from complementary disciplines, the international PLANTCOM network identified four timely questions to foster a better understanding of the mechanisms mediating plant assemblages. We propose that by identifying the key relationships among phenotypic traits involved in plant-plant interactions and the underlying adaptive genetic and molecular pathways, while considering environmental fluctuations at diverse spatial and time scales, we can improve predictions of genotype-by-genotype-by-environment interactions and modeling of productive and stable plant assemblages in wild habitats and crop fields.

Seed dormancy is a crucial developmental transition that affects the adaption and survival of plants. Arabidopsis DELAY OF GERMINATION 1 (DOG1) is known as a master regulator of seed dormancy. However, although several upstream factors of DOG1 have been reported, the exact regulation of DOG1 is not fully understood. Histone acetylation is an important regulatory layer, controlled by histone acetyltransferases and histone deacetylases. Histone acetylation strongly correlates with transcriptionally active chromatin, whereas heterochromatin is generally characterized by hypoacetylated histones. Here we describe that loss of function of two plant-specific histone deacetylases, HD2A and HD2B, resulted in enhanced seed dormancy in Arabidopsis. Interestingly, the silencing of HD2A and HD2B caused hyperacetylation of the DOG1 locus and promoted the expression of DOG1 during seed maturation and imbibition. Knockout of DOG1 could rescue the seed dormancy and partly rescue the disturbed development phenotype of hd2ahd2b. Transcriptomic analysis of the hd2ahd2b line shows that many genes involved in seed development were impaired. Moreover, we demonstrated that HSI2 and HSL1 interact with HD2A and HD2B. In sum, these results suggest that HSI2 and HSL1 might recruit HD2A and HD2B to DOG1 to negatively regulate DOG1 expression and to reduce seed dormancy, consequently, affecting seed development during seed maturation and promoting seed germination during imbibition.

Genome evolution is partly driven by the mobility of transposable elements (TEs) which often leads to deleterious effects, but their activity can also facilitate genetic novelty and catalyze local adaptation. We explored how the intraspecific diversity of TE polymorphisms is shaping the broad geographic success and adaptation capacity of the emerging oil cropThlaspi arvense. We achieved this by classifying the TE inventory of this species based on a high-quality genome assembly, age estimation of retrotransposon TE families and a comprehensive assessment of their mobilization potential. Our survey of TE insertion polymorphisms (TIPs) captured 280 accessions from 12 regions across the Northern hemisphere. We quantified over 90,000 TIPs, with their distribution mirroring genetic differentiation as measured by single nucleotide polymorphisms (SNPs). The number and types of mobile TE families vary substantially across populations, but there are also shared patterns common to all accessions. We found that Ty3/Athila elements are the main drivers of TE diversity inT. arvensepopulations, while a single Ty1/Alesia lineage might be particularly important for molding transcriptome divergence. We further observed that the number of retrotransposon TIPs is associated with variation at genes related to epigenetic regulation while DNA transposons are associated with variation at a Heat Shock Protein (HSP19). We propose that the high rate of mobilization activity can be harnessed for targeted gene expression diversification, which may ultimately present a toolbox for the potential use of transposition in breeding and domestication ofT. arvense.

Due to the accelerating climate change, it is crucial to understand how plants adapt to rapid environmental changes. Such adaptation may be mediated by epigenetic mechanisms like DNA methylation, which could heritably alter phenotypes without changing the DNA sequence, especially across clonal generations. However, we are still missing robust evidence of the adaptive potential of DNA methylation in wild clonal populations. Here, we studied the methylome of Fragaria vesca, a predominantly clonally reproducing herb. We examined samples from 21 natural populations across three climatically distinct geographic regions, as well as clones of the same individuals grown in a common garden. We found that inherited epigenetic variation was partly associated with climate of origin, and that a subset of these epigenetic changes affected gene expression. Our results indicate that DNA methylation variation in the wild is common and may contribute to adaptation of clonal plant populations.

Since the first half of the twentieth century, evolutionary theory has been dominated by the idea that mutations occur randomly with respect to their consequences1. Here we test this assumption with large surveys of de novo mutations in the plant Arabidopsis thaliana. In contrast to expectations, we find that mutations occur less often in functionally constrained regions of the genome—mutation frequency is reduced by half inside gene bodies and by two-thirds in essential genes. With independent genomic mutation datasets, including from the largest Arabidopsis mutation accumulation experiment conducted to date, we demonstrate that epigenomic and physical features explain over 90% of variance in the genome-wide pattern of mutation bias surrounding genes. Observed mutation frequencies around genes in turn accurately predict patterns of genetic polymorphisms in natural Arabidopsis accessions (r = 0.96). That mutation bias is the primary force behind patterns of sequence evolution around genes in natural accessions is supported by analyses of allele frequencies. Finally, we find that genes subject to stronger purifying selection have a lower mutation rate. We conclude that epigenome-associated mutation bias2 reduces the occurrence of deleterious mutations in Arabidopsis, challenging the prevailing paradigm that mutation is a directionless force in evolution.

Summary- Environmental changes can trigger phenotypic variation in plants through epigenetic mechanisms, but strong genetic influences make it difficult to isolate and study epigenetic effects. Clonal trees with low genetic variation, such as the Lombardy poplar(Populus nigracv. ‘Italica’ Duroi), offer a unique system to study epigenetic variation associated with the environment.- We collected cuttings (ramets) of Lombardy poplar along a wide geographical range in Europe. We performed whole-genome-bisulfite sequencing of 164 ramets grown in a common garden and of a subset of 35 of the original parental individuals. Using historical bioclimatic data, we tested the relationship between DNA methylation and climatic gradients.- We found that average methylation levels in TEs and promoter regions correlate with biologically relevant climatic variables. Furthermore, we observed that DNA methylation was transmitted to the next clonal generation, but a fraction of the methylome changed relatively fast when comparing the parental individuals with the clonal offspring.- Our results suggest that the poplar methylome is a dynamic layer of information that can be transmitted to the clonal offspring and potentially affect how poplars acclimate to new environmental conditions.

Background Plants growing in proximity to other plants are exposed to a variety of metabolites that these neighbors release into the environment. Some species produce allelochemicals to inhibit growth of neighboring plants, which in turn have evolved ways to detoxify these compounds. Results In order to understand how the allelochemical-receiving target plants respond to chemically diverse compounds, we performed whole-genome transcriptome analysis of Arabidopsis thaliana exposed to either the benzoxazinoid derivative 2-amino- 3H-phenoxazin-3-one (APO) or momilactone B. These two allelochemicals belong to two very different compound classes, benzoxazinoids and diterpenes, respectively, produced by different Poaceae crop species. Conclusions Despite their distinct chemical nature, we observed similar molecular responses of A. thaliana to these allelochemicals. In particular, many of the same or closely related genes belonging to the three-phase detoxification pathway were upregulated in both treatments. Further, we observed an overlap between genes upregulated by allelochemicals and those involved in herbicide detoxification. Our findings highlight the overlap in the transcriptional response of a target plant to natural and synthetic phytotoxic compounds and illustrate how herbicide resistance could arise via pathways involved in plant-plant interaction.

This repository stores data produced from a genomic and epigenomic large-scale survey of natural populations of Thlaspi arvense, investigating the genetic and environmental bases of epigenetic variation. 207 lines from across Europe were grown in common environment and scored for genetic and DNA methylation variation by Whole Genome Sequencing and Whole Genome Bisulfite Sequencing respectively. This repository includes: 1) The original mean methylation values for all lines, the respective processed values (corrected for non-conversion rate by subtracting the "Surplus_0.6NCR", and optionally corrected for coverage and transformed), used for Genome Wide Association Studies (GWAS) and correlation with climatic variables. In addition, the weighted methylation values (Schultz et al. 2012) are available. 2) Differentially Methylated Regions and Variance Decomposition Analysis results, obtained with scripts available at https://github.com/Dario-Galanti/popDMRs_refine_VCA. 3) Gene Body Methylation (GBM) classification for each line and sequence context. A binomial test was applied to each coding sequence (CDS) to determine if its fraction of methylated cytosines was higher than average (Takuno and Gaut 2013). A 0 indicates a non-significant test, a 1 indicates a significant test. Code available at https://github.com/Dario-Galanti/WGBS_downstream/tree/main/WGBS_completeworkflow. 4) GWAS results (filtered for -log(p)>1) for all processed mean methylation values are available with the ".gwas" extention and can be opened with Integrative Genomics Viewer (https://software.broadinstitute.org/software/igv/), using the reference genome published by Nunn et al. 2021 (https://pubmed.ncbi.nlm.nih.gov/34990041/)., {"references":["Schultz MD, Schmitz RJ, Ecker JR. 'Leveling' the playing field for analyses of single-base resolution DNA methylomes. Trends Genet. 2012 Dec 1;28(12):583–5","Takuno S, Gaut BS. Gene body methylation is conserved between plant orthologs and is of evolutionary consequence. Proc Natl Acad Sci. 2013 Jan 29;110(5):1797–802","Nunn A, Rodríguez-Arévalo I, Tandukar Z, Frels K, Contreras-Garrido A, Carbonell-Bejerano P, et al. Chromosome-level Thlaspi arvense genome provides new tools for translational research and for a newly domesticated cash cover crop of the cooler climates. Plant Biotechnology Journal. 2022 Aug; 1–20. Available from: https://onlinelibrary.wiley.com/doi/full/10.1111/pbi.13775"]}

Clonal propagation is frequently used in commercial plant breeding and biotechnology programs because it minimizes genetic variation, yet it is not uncommon to observe clonal plants with stable phenotypic changes, a phenomenon known as somaclonal variation. Several studies have linked epigenetic modifications induced during regeneration with this newly acquired phenotypic variation. However, the factors that determine the extent of somaclonal variation and the molecular changes underpinning this process remain poorly understood. To address this gap in our knowledge, we compared clonally propagated Arabidopsis thaliana plants derived from somatic embryogenesis using two different embryonic transcription factors- RWP-RK DOMAIN-CONTAINING 4 (RKD4) or LEAFY COTYLEDON2 (LEC2) and from two epigenetically distinct founder tissues. We found that both the epi(genetic) status of the explant and the regeneration protocol employed play critical roles in shaping the molecular and phenotypic landscape of clonal plants. Phenotypic variation in regenerated plants can be largely explained by the inheritance of tissue-specific DNA methylation imprints, which are associated with specific transcriptional and metabolic changes in sexual progeny of clonal plants. For instance, regenerants were particularly affected by the inheritance of root-specific epigenetic imprints, which were associated with an increased accumulation of salicylic acid in leaves and accelerated plant senescence. Collectively, our data reveal specific pathways underpinning the phenotypic and molecular variation that arise and accumulate in clonal plant populations.

Environmental stresses can result in a wide range of physiological and molecular responses in plants. These responses can also impact epigenetic information in genomes, especially at the level of DNA methylation (5-methylcytosine). DNA methylation is the hallmark heritable epigenetic modification and plays a key role in silencing transposable elements (TEs). Although DNA methylation is an essential epigenetic mechanism, fundamental aspects of its contribution to stress responses and adaptation remain obscure. We investigated epigenome dynamics of wild strawberry (

Although clonal propagation is frequently used in commercial plant breeding and plant biotechnology programs because it minimizes genetic variation, it is not uncommon to observe clonal plants with stable phenotypic changes, a phenomenon known as somaclonal variation. Several studies have shown that epigenetic modifications induced during regeneration are associated with this newly acquired phenotypic variation. However, the factors that determine the extent of somaclonal variation and the molecular changes associated with it remain poorly understood. To address this gap in our knowledge, we compared clonally propagated Arabidopsis thaliana plants derived from somatic embryogenesis using two different embryonic transcription factors-RWP-RK DOMAIN-CONTAINING 4 (RKD4) or LEAFY COTYLEDON2 (LEC2) and from two epigenetically distinct tissues. We found that both the epi(genetic) status of explant and the regeneration protocol employed play critical roles in shaping the molecular and phenotypic state of clonal plants. Phenotypic variation of regenerated plants can be largely explained by the inheritance of tissue-specific DNA methylation imprints, which are associated with specific transcriptional and metabolic changes in sexual progeny of clonal plants. Moreover, regenerants from roots were particularly affected by the inheritance of epigenetic imprints, which resulted in increased accumulation of salicylic acid in leaves and accelerated plant senescence. Collectively, our data reveal pathways for targeted manipulation of phenotypic variation in clonal plants.

SummaryEnvironmental stresses can result in a wide range of physiological and molecular responses in plants. These responses can also impact epigenetic information in genomes especially at the level of DNA methylation. DNA methylation is the hallmark heritable epigenetic modification and plays a key role in silencing transposable elements (TEs). Although DNA methylation is an essential epigenetic mechanism, fundamental aspects of its contribution to stress responses and adaptation remain obscure.We investigated epigenome dynamics of wild strawberry (Fragaria vesca) in response to variable environmental conditions at DNA methylation level. F. vesca methylome responded with great plasticity to ecologically relevant abiotic and hormonal stresses. Thermal stress resulted in substantial genome-wide loss of DNA methylation. Notably, all tested stress conditions resulted in marked hot spots of differential DNA methylation near centromeric or pericentromeric regions, particularly in non-symmetrical DNA methylation context. Additionally, we identified differentially methylated regions (DMRs) within promoter regions of transcription factor (TF) superfamilies involved in plant stress-response and assessed the effects of these changes on gene expression.These findings improve our understanding on stress-response at the epigenome level by highlighting the correlation between DNA methylation, TEs and gene expression regulation in plants subjected to a broad range of environmental stresses.

Allelopathy is a biochemical interaction between plants in which a donor plant releases secondary metabolites, allelochemicals, that are detrimental to the growth of its neighbours. Traditionally considered as bilateral interactions between two plants, allelopathy has recently emerged as a cross-kingdom process that can influence and be modulated by the other organisms in the plant's environment. Here, we review the current knowledge on plant-interkingdom interactions, with a particular focus on benzoxazinoids. We highlight how allelochemical-producing plants influence not only their plant neighbours but also insects, fungi, and bacteria that live on or around them. We discuss challenges that need to be overcome to study chemical plant-interkingdom interactions, and we propose experimental approaches to address how biotic and chemical processes impact plant health.

Summary Environmental stresses can result in a wide range of physiological and molecular responses in plants. These responses can also impact epigenetic information in genomes, especially at the level of DNA methylation (5-methylcytosine). DNA methylation is the hallmark heritable epigenetic modification and plays a key role in silencing transposable elements (TEs). Although DNA methylation is an essential epigenetic mechanism, fundamental aspects of its contribution to stress responses and adaptation remain obscure. We investigated epigenome dynamics of wild strawberry (Fragaria vesca) in response to variable ecologically relevant environmental conditions at the DNA methylation level. F. vesca methylome responded with great plasticity to ecologically relevant abiotic and hormonal stresses. Thermal stress resulted in substantial genome-wide loss of DNA methylation. Notably, all tested stress conditions resulted in marked hot spots of differential DNA methylation near centromeric or pericentromeric regions, particularly in the non-symmetrical DNA methylation context. Additionally, we identified differentially methylated regions (DMRs) within promoter regions of transcription factor (TF) superfamilies involved in plant stress-response and assessed the effects of these changes on gene expression. These findings improve our understanding on stress-response at the epigenome level by highlighting the correlation between DNA methylation, TEs and gene expression regulation in plants subjected to a broad range of environmental stresses.

Whole-genome bisulfite sequencing (WGBS) is the standard method for profiling DNA methylation at single-nucleotide resolution. Different tools have been developed to extract differentially methylated regions (DMRs), often built upon assumptions from mammalian data. Here, we present MethylScore, a pipeline to analyse WGBS data and to account for the substantially more complex and variable nature of plant DNA methylation. MethylScore uses an unsupervised machine learning approach to segment the genome by classification into states of high and low methylation. It processes data from genomic alignments to DMR output and is designed to be usable by novice and expert users alike. We show how MethylScore can identify DMRs from hundreds of samples and how its data-driven approach can stratify associated samples without prior information. We identify DMRs in the A. thaliana 1,001 Genomes dataset to unveil known and unknown genotype–epigenotype associations .

Whole-genome bisulfite sequencing (WGBS) is the standard method for profiling DNA methylation at single-nucleotide resolution. Many WGBS-based studies aim to identify biologically relevant loci that display differential methylation between genotypes, treatment groups, tissues, or developmental stages. Over the years, different tools have been developed to extract differentially methylated regions (DMRs) from whole-genome data. Often, such tools are built upon assumptions from mammalian data and do not consider the substantially more complex and variable nature of plant DNA methylation. Here, we present MethylScore, a pipeline to analyze WGBS data and to account for plant-specific DNA methylation properties. MethylScore processes data from genomic alignments to DMR output and is designed to be usable by novice and expert users alike. It uses an unsupervised machine learning approach to segment the genome by classification into states of high and low methylation, substantially reducing the number of necessary statistical tests while increasing the signal-to-noise ratio and the statistical power. We show how MethylScore can identify DMRs from hundreds of samples and how its data-driven approach can stratify associated samples without prior information. We identify DMRs in the A. thaliana 1001 Genomes dataset to unveil known and unknown genotype-epigenotype associations. MethylScore is an accessible pipeline for plant WGBS data, with unprecedented features for DMR calling in small- and large-scale datasets; it is built as a Nextflow pipeline and its source code is available at https://github.com/Computomics/MethylScore.

Low-free-energy surfaces have attracted an intense academic and industrial interest over the last decade. A reduction of the surface free energy (SFE) has been found to enhance self-cleaning, hydrophobic, and non-fouling properties of surfaces, which are highly desirable in many industrial applications. However, tuning the surface chemistry and topography to achieve tailored low free energy surfaces has been found extremely challenging. In this work, we first show that an accurate refinement of the atmospheric plasma technique guarantees a polymeric coating near to the super-hydrophobic regime. Second, by coupling modelling and experimental measurements we suggest a reliable workflow for the surface characterization and smart design. Specifically, the case study proposed in this contest is capable of quantitatively distinguishing the contribution of a Perfluoro Decyl Acrylate (PFDA) coating, and hence decoupling the role of surface chemistry and roughness, in the reduction of the surface free energy of a bare silicon sample. Beyond the specific case study, our results also emphasize that a synergistic combination of models and experiments can unveil the optimal pathway for designing low-free-energy surfaces.

Thlaspi arvense (field pennycress) is being domesticated as a winter annual oilseed crop capable of improving ecosystems and intensifying agricultural productivity without increasing land use. It is a selfing diploid with a short life cycle and is amenable to genetic manipulations, making it an accessible field-based model species for genetics and epigenetics. The availability of a high quality reference genome is vital for understanding pennycress physiology and for clarifying its evolutionary history within the Brassicaceae. Here, we present a chromosome-level genome assembly of var. MN106-Ref with improved gene annotation, and use it to investigate gene structure differences between two accessions (MN108 and Spring32-10) that are highly amenable to genetic transformation. We describe small RNAs, pseudogenes, and transposable elements, and highlight tissue specific expression and methylation patterns. Resequencing of forty wild accessions provides insights into genome-wide genetic variation as well as QTL regions for flowering time and a seedling color phenotype. Altogether, these data will serve as a tool for pennycress improvement in general and for translational research across the Brassicaceae.

Linking plant phenotype to genotype is a common goal to both plant breeders and geneticists. However, collecting phenotypic data for large numbers of plants remain a bottleneck. Plant phenotyping is mostly image based and therefore requires rapid and robust extraction of phenotypic measurements from image data. However, because segmentation tools usually rely on color information, they are sensitive to background or plant color deviations. We have developed a versatile, fully open-source pipeline to extract phenotypic measurements from plant images in an unsupervised manner. ARADEEPOPSIS (https://github.com/Gregor-Mendel-Institute/aradeepopsis) uses semantic segmentation of top-view images to classify leaf tissue into three categories: healthy, anthocyanin rich, and senescent. This makes it particularly powerful at quantitative phenotyping of different developmental stages, mutants with aberrant leaf color and/or phenotype, and plants growing in stressful conditions. On a panel of 210 natural Arabidopsis (Arabidopsis thaliana) accessions, we were able to not only accurately segment images of phenotypically diverse genotypes but also to identify known loci related to anthocyanin production and early necrosis in genome-wide association analyses. Our pipeline accurately processed images of diverse origin, quality, and background composition, and of a distantly related Brassicaceae. ARADEEPOPSIS is deployable on most operating systems and high-performance computing environments and can be used independently of bioinformatics expertise and resources.

In the past, reactive nitrogen species (RNS) were supposed to be stress-induced by-products of disturbed metabolism that cause oxidative damage to biomolecules. However, emerging evidence demonstrates a substantial role of RNS as endogenous signals in eukaryotes. In plants, S-nitrosoglutathione (GSNO) is the dominant RNS and serves as the •NO donor for S-nitrosation of diverse effector proteins. Remarkably, the endogenous GSNO level is tightly controlled by S-nitrosoglutathione reductase (GSNOR) that irreversibly inactivates the glutathione-bound NO to ammonium. Exogenous feeding of diverse RNS, including GSNO, affected chromatin accessibility and transcription of stress-related genes, but the triggering function of RNS on these regulatory processes remained elusive. Here, we show that GSNO reductase-deficient plants (gsnor1-3) accumulate S-adenosylmethionine (SAM), the principal methyl donor for methylation of DNA and histones. This SAM accumulation triggered a substantial increase in the methylation index (MI = [SAM]/[S-adenosylhomocysteine]), indicating the transmethylation activity and histone methylation status in higher eukaryotes. Indeed, a mass spectrometry-based global histone profiling approach demonstrated a significant global increase in H3K9me2, which was independently verified by immunological detection using a selective antibody. Since H3K9me2-modified regions tightly correlate with methylated DNA regions, we also determined the DNA methylation status of gsnor1-3 plants by whole-genome bisulfite sequencing. DNA methylation in the CG, CHG, and CHH contexts in gsnor1-3 was significantly enhanced compared to the wild type. We propose that GSNOR1 activity affects chromatin accessibility by controlling the transmethylation activity (MI) required for maintaining DNA methylation and the level of the repressive chromatin mark H3K9me2.

Plants are hubs of organismic interactions. They constantly engage in beneficial or competitive interactions with fungi, oomycetes, bacteria, insects, nematodes, and other plants. To adjust the molecular processes necessary for the establishment and maintenance of beneficial interactions and for the defense against pathogens and herbivores, plants have evolved intricate regulatory mechanisms. Besides the canonical plant immune system that acts as the primary defense, epigenetic mechanisms have started to emerge as another regulatory entity and as a target of pathogens trying to overcome the plant’s defenses. In this review, we highlight recent advances in understanding the contribution of various epigenetic components and of epigenetic diversity to plantorganismic interactions.

Objective: To evaluate the immunogenicity, safety, and efficacy of etanercept (ETN) manufactured using the serum-free, high-capacity manufacturing (SFHCM) process in patients with rheumatoid arthritis (RA). ----- Methods: In this global, multicenter, open-label, single-arm study (NCT02378506), 187 adult patients with moderate to severe RA received ETN 50 mg once weekly for 24 weeks manufactured using the SFHCM process. Immunogenicity (presence of antidrug antibodies (ADAs) and neutralizing antibodies (NAbs)) was assessed at 12 and 24 weeks. Safety and efficacy were evaluated at 4, 12, and 24 weeks. ----- Results: Eight (4.5%) patients tested positive for ADA, and there were no NAbs detected at any time throughout the study. Ninety (48.1%) patients reported treatment-emergent adverse events (AEs), of which 27 (14.4%) reported injection-site reactions, and 43 (23.0%) reported infections. The majority of AEs were mild or moderate in severity, and the drug was well tolerated. Throughout the duration of the study (week 4 to week 24), there was a progressive increase in the American College of Rheumatology (ACR)-defined responses (ACR20: 55.9%–82.0%, ACR50: 16.1%–57.8%, and ACR70: 3.2%–26.7%) from baseline and the proportion of patients achieving low disease activity and remission, with a corresponding decrease in measures of disease activity. ----- Conclusion: The immunogenicity, safety, and efficacy of ETN manufactured using the SFHCM process were similar to the current approved ETN formulation. ClinicalTrials.gov registration: NCT02378506.

Bisulfite sequencing is a widely used technique for determining DNA methylation and its relationship with epigenetics, genetics, and environmental parameters. Various techniques were implemented for epigenome-wide association studies (EWAS) to reveal meaningful associations, however, there are only very few plant studies available to date. Here, we developed the EpiDiverse EWAS pipeline and tested it using two plant datasets, from P. abies (Norway spruce) and Q. lobata (valley oak). Hence, we present an EWAS implementation tested for non-model plant species and describe its use.

Nitric oxide (NO) is a signaling molecule with multiple regulatory functions in plant physiology and stress response. In addition to direct effects on transcriptional machinery, NO executes its signaling function via epigenetic mechanisms. We report that light intensity-dependent changes in NO correspond to changes in global histone acetylation (H3, H3K9, and H3K9/K14) in Arabidopsis (Arabidopsis thaliana) wild-type leaves, and that this relationship depends on S-nitrosoglutathione reductase and histone deacetylase 6 (HDA6). The activity of HDA6 was sensitive to NO, demonstrating that NO participates in regulation of histone acetylation. Chromatin immunoprecipitation sequencing and RNA-seq analyses revealed that NO participates in the metabolic switch from growth and development to stress response. This coordinating function of NO might be particularly important in plant ability to adapt to a changing environment, and is therefore a promising foundation for mitigating the negative effects of climate change on plant productivity.

DNA methylation has evolved to silence mutagenic transposable elements (TEs) while typically avoiding the targeting of endogenous genes. Mechanisms that prevent DNA methyltransferases from ectopically methylating genes are expected to be of prime importance during periods of dynamic cell cycle activities including plant embryogenesis. However, virtually nothing is known regarding how DNA methyltransferase activities are precisely regulated during embryogenesis to prevent the induction of potentially deleterious and mitotically stable genic epimutations. Here, we report that microRNA-mediated repression of CHROMOMETHYLASE 3 (CMT3) and the chromatin features that CMT3 prefers help prevent ectopic methylation of thousands of genes during embryogenesis that can persist for weeks afterwards. Our results are also consistent with CMT3-induced ectopic methylation of promoters or bodies of genes undergoing transcriptional activation reducing their expression. Therefore, the repression of CMT3 prevents epigenetic collateral damage on endogenous genes. We also provide a model that may help reconcile conflicting viewpoints regarding the functions of gene-body methylation that occurs in nearly all flowering plants.

In this research a high-throughput RNA sequencing based transcriptome analysis technique (RNA-Seq) was used to evaluate differentially expressed genes (DEGs) in the wild type Arabidopsis seedling in response to flg22, a well-known microbe-associated molecular pattern (MAMP), and AtPep1, a well-known peptide representing an endogenous damage-associated molecular patterns (DAMP). The results of our study revealed that 1895 (1634 up-regulated and 261 down-regulated) and 2271 (1706 up-regulated and 565 down-regulated) significant differentially expressed genes in response to flg22 and AtPep1 treatment, respectively. Among significant DEGs, we observed that a number of hitherto overlooked genes have been found to be induced upon treatment with either flg22 or with AtPep1, indicating their possible involvement in innate immunity. Here, we characterized two of them, namely PP2-B13 and ACLP1. pp2-b13 and aclp1 mutants showed an increased susceptibility to infection by the virulent pathogen Pseudomomas syringae pv tomato mutant hrcC-, as evidenced by an increased growth of the pathogen in planta. Further we present evidence that the aclp1 mutant was deficient in ethylene production upon flg22 treatment, while the pp2-b13 mutant, was deficient in ROS production. The results from this research provide new information to a better understanding of the immune system in Arabidopsis.

DNA methylation has evolved to silence mutagenic transposable elements (TEs) while typically avoiding the targeting of endogenous genes. Mechanisms that prevent DNA methyltransferases from ectopically methylating genes are expected to be of prime importance during periods of dynamic cell cycle activities including plant embryogenesis. However, virtually nothing is known regarding how DNA methyltransferase activities are precisely regulated during embryogenesis to prevent the induction of potentially deleterious and mitotically stable genic epimutations. Here, we report that microRNA-mediated repression of CHROMOMETHYLASE 3 (CMT3) and the chromatin features that CMT3 prefers help prevent ectopic methylation of thousands of genes during embryogenesis that can persist for weeks afterwards. Moreover, CMT3-induced ectopic methylation of genes undergoing transcriptional activation can reduce their corresponding transcript levels. Therefore, the repression of CMT3 prevents epigenetic collateral damage on endogenous genes. We also provide a model that may help reconcile conflicting viewpoints regarding the functions of gene-body methylation that occurs in nearly all flowering plants.

In the struggle to secure nutrient access and to outperform competitors, some plant species have evolved a biochemical arsenal with which they inhibit the growth or development of neighbouring plants. This process, known as allelopathy, exists in many of today’s major crops, including rice. Rice synthesizes momilactones, diterpenoids that are released into the rhizosphere and inhibit the growth of numerous plant species. While the allelopathic potential of rice was recognized decades ago, many questions remain unresolved regarding the biosynthesis, exudation, and biological activity of momilactones. Here, we review current knowledge on momilactones, their role in allelopathy, and their potential to serve as a basis for sustainable weed management. We emphasize the gaps in our current understanding of when and how momilactones are produced and of how they act in plant cells, and outline what we consider the next steps in momilactone and rice allelopathy research.

Nitric oxide (NO) is a signaling molecule with multiple regulatory functions in plant physiology and stress response. Besides direct effects on the transcriptional machinery, NO can fulfill its signaling function via epigenetic mechanisms.We report that light intensity-dependent changes in NO correlate with changes in global histone acetylation (H3, H3K9 and H3K9/K14) in Arabidopsis thaliana wild-type leaves and that this correlation depends on S-nitrosoglutathione reductase and histone deacetylase 6. The activity of histone deacetylase 6 was sensitive to NO, which demonstrates that NO participates in regulation of histone acetylation. ChIP-seq and RNA-seq analyses revealed that NO is involved in the metabolic switch from growth and development to stress response. This coordinating function of NO might be of special importance in adaptation to a changing environment and could therefore be a promising starting point to mitigating the negative effects of climate change on plant productivity.

Linking plant phenotype to genotype, i.e., identifying genetic determinants of phenotypic traits, is a common goal of both plant breeders and geneticists. While the ever-growing genomic resources and rapid decrease of sequencing costs have led to enormous amounts of genomic data, collecting phenotypic data for large numbers of plants remains a bottleneck. Many phenotyping strategies rely on imaging plants, which makes it necessary to extract phenotypic measurements from these images rapidly and robustly. Common image segmentation tools for plant phenotyping mostly rely on color information, which is error-prone when either background or plant color deviate from the underlying expectations. We have developed a versatile, fully open-source pipeline to extract phenotypic measurements from plant images in an unsupervised manner. aradeepopsis was built around the deep-learning model DeepLabV3+ that was re-trained for segmentation of Arabidopsis thaliana rosettes. It uses semantic segmentation to classify leaf tissue into up to three categories: healthy, anthocyanin-rich, and senescent. This makes aradeepopsis particularly powerful at quantitative phenotyping from early to late developmental stages, of mutants with aberrant leaf color and/or phenotype, and of plants growing in stressful conditions where leaf color may deviate from green. Using our tool on a panel of 210 natural Arabidopsis accessions, we were able to not only accurately segment images of phenotypically diverse genotypes but also to map known loci related to anthocyanin production and early necrosis using the aradeepopsis output in genome-wide association analyses. Our pipeline is able to handle images of diverse origins, image quality, and background composition, and could even accurately segment images of a distantly related Brassicaceae. Because it can be deployed on virtually any common operating system and is compatible with several high-performance computing environments, aradeepopsis can be used independently of bioinformatics expertise and computing resources. aradeepopsis is available at https://github.com/Gregor-Mendel-Institute/aradeepopsis.

Studies on "epigenetic inheritance" or "transgenerational epigenetic inheritance" have emerged at ever-increasing numbers in the last years, in plant as well as animal systems and in diverse contexts ranging from stress adaptation to behavioral studies. Despite the very different organisms and biological processes investigated, the overarching question has been if and how an organism's epigenome registers and records external cues and relays this information to its progeny. Very often, these experiments are based on the hypothesis that epigenetic memorization of events or conditions could be the basis of an altered response of the progeny upon encountering the same or a similar condition. If confirming the hypothesis, such studies challenge our fundamental understanding of evolution by natural selection; therefore they require particular rigor in design and great care in data analysis. Here, we want to provide general guidelines on how to design studies on epigenetic inheritance in plants and to consider critical points during data analysis and interpretation. While we cannot provide a step-by-step guide that fits all experimental setups and questions addressed, we explain frequent misconceptions and often overlooked pitfalls. Our aim is to provide researchers with conceptual tools to sensibly design their studies and to interpret their results in the admissible framework.

Linking plant phenotype to genotype is a common goal to both plant breeders and geneticists. However, collecting phenotypic data for large numbers of plants remain a bottleneck. Plant phenotyping is mostly image based and therefore requires rapid and robust extraction of phenotypic measurements from image data. However, because segmentation tools usually rely on color information, they are sensitive to background or plant color deviations. We have developed a versatile, fully open-source pipeline to extract phenotypic measurements from plant images in an unsupervised manner. ARADEEPOPSIS (https://github.com/Gregor-Mendel-Institute/aradeepopsis) uses semantic segmentation of top-view images to classify leaf tissue into three categories: healthy, anthocyanin rich, and senescent. This makes it particularly powerful at quantitative phenotyping of different developmental stages, mutants with aberrant leaf color and/or phenotype, and plants growing in stressful conditions. On a panel of 210 natural Arabidopsis (Arabidopsis thaliana) accessions, we were able to not only accurately segment images of phenotypically diverse genotypes but also to identify known loci related to anthocyanin production and early necrosis in genome-wide association analyses. Our pipeline accurately processed images of diverse origin, quality, and background composition, and of a distantly related Brassicaceae. ARADEEPOPSIS is deployable on most operating systems and high-performance computing environments and can be used independently of bioinformatics expertise and resources.

Studies on “epigenetic inheritance” or “transgenerational epigenetic inheritance” have emerged at ever-increasing numbers in the last years, in plant as well as animal systems and in diverse contexts ranging from stress adaptation to behavioral studies. Despite the very different organisms and biological processes investigated, the overarching question has been if and how an organism’s epigenome registers and records external cues and relays this information to its progeny. Very often, these experiments are based on the hypothesis that epigenetic memorization of events or conditions could be the basis of an altered response of the progeny upon encountering the same or a similar condition. If confirming the hypothesis, such studies challenge our fundamental understanding of evolution by natural selection; therefore they require particular rigor in design and great care in data analysis. Here, we want to provide general guidelines on how to design studies on epigenetic inheritance in plants and to consider critical points during data analysis and interpretation. While we cannot provide a step-by-step guide that fits all experimental setups and questions addressed, we explain frequent misconceptions and often overlooked pitfalls. Our aim is to provide researchers with conceptual tools to sensibly design their studies and to interpret their results in the admissible framework.

Summary Plant‐specific EFFECTORS OF TRANSCRIPTION (ET) are characterised by a variable number of highly conserved ET repeats, which are involved in zinc and DNA binding. In addition, ETs share a GIY‐YIG domain, involved in DNA nicking activity. It was hypothesised that ETs might act as epigenetic regulators.Here, methylome, transcriptome and phenotypic analyses were performed to investigate the role of ET factors and their involvement in DNA methylation in Arabidopsis thaliana.Comparative DNA methylation and transcriptome analyses in flowers and seedlings of et mutants revealed ET‐specific differentially expressed genes and mostly independently characteristic, ET‐specific differentially methylated regions. Loss of ET function results in pleiotropic developmental defects.The accumulation of cyclobutane pyrimidine dimers after ultraviolet stress in et mutants suggests an ET function in DNA repair.

The epigenome orchestrates genome accessibility, functionality, and three-dimensional structure. Because epigenetic variation can impact transcription and thus phenotypes, it may contribute to adaptation. Here, we report 1,107 high-quality single-base resolution methylomes and 1,203 transcriptomes from the 1001 Genomes collection of Arabidopsis thaliana. Although the genetic basis of methylation variation is highly complex, geographic origin is a major predictor of genome-wide DNA methylation levels and of altered gene expression caused by epialleles. Comparison to cistrome and epicistrome datasets identifies associations between transcription factor binding sites, methylation, nucleotide variation, and co-expression modules. Physical maps for nine of the most diverse genomes reveal how transposons and other structural variants shape the epigenome, with dramatic effects on immunity genes. The 1001 Epigenomes Project provides a comprehensive resource for understanding how variation in DNA methylation contributes to molecular and non-molecular phenotypes in natural populations of the most studied model plant.

Summary Auxin gradients are sustained by series of influx and efflux carriers whose subcellular localization is sensitive to both exogenous and endogenous factors. Recently the localization of the Arabidopsis thaliana auxin efflux carrier PIN-FORMED (PIN) 6 was reported to be tissue-specific and regulated through unknown mechanisms. Here, we used genetic, molecular and pharmacological approaches to characterize the molecular mechanism(s) controlling the subcellular localization of PIN6. PIN6 localizes to endomembrane domains in tissues with low PIN6 expression levels such as roots, but localizes at the plasma membrane (PM) in tissues with increased PIN6 expression such as the inflorescence stem and nectary glands. We provide evidence that this dual localization is controlled by PIN6 phosphorylation and demonstrate that PIN6 is phosphorylated by mitogen-activated protein kinases (MAPKs) MPK4 and MPK6. The analysis of transgenic plants expressing PIN6 at PM or in endomembrane domains reveals that PIN6 subcellular localization is critical for Arabidopsis inflorescence stem elongation post-flowering (bolting). In line with a role for PIN6 in plant bolting, inflorescence stems elongate faster in pin6 mutant plants than in wild-type plants. We propose that PIN6 subcellular localization is under the control of developmental signals acting on tissue-specific determinants controlling PIN6-expression levels and PIN6 phosphorylation.

Transposable elements (TEs) are prevalent in most eukaryotes, and host genomes have devised silencing strategies to rein in TE activity. One of these, transcriptional silencing, is generally associated with DNA methylation and short interfering RNAs. Here we show that the Arabidopsis genes MAIL1 and MAIN define an alternative silencing pathway independent of DNA methylation and short interfering RNAs. Mutants for MAIL1 or MAIN exhibit release of silencing and appear to show impaired condensation of pericentromeric heterochromatin. Phylogenetic analysis suggests not only that MAIL1 and MAIN encode a retrotransposon-related plant mobile domain, but also that host plant mobile domains were captured by DNA transposons during plant evolution. Our results reveal a role for Arabidopsis proteins with a transposon-related domain in gene silencing., Host genomes have evolved multiple silencing mechanisms to repress transposable element activity. Here Ikeda et al. show that the Arabidopsis MAIL1 and MAIN genes encode a retrotransposon-related plant mobile domain and define an alternative silencing pathway largely independent of DNA methylation and siRNAs.

Epigenetic variation - polymorphisms at the level of DNA methylation or histone modifications - modulates chromatin accessibility, which can perturb transcriptional activity and spur phenotypic variation. Determining the origin, frequency spectrum, and consequences of epigenetic variants is key to understanding the role of this variation in generating stable phenotypic variation in plants. Here we review recent literature on DNA methylation variation in both model and crop plant species with a focus on the link between genotype, epigenotype, and transcription. We highlight population epigenomics studies that explore the relationship between epigenetic variants and genetic diversity. Moreover, we provide an overview of relevant studies that together advocate a minor, albeit significant role for epigenetic variation in directing specific transcriptional changes.

Across the plant kingdom, phytochrome (PHY) photoreceptors play an important role during adaptive and developmental responses to light. In Arabidopsis thaliana, light-activated PHYs accumulate in the nucleus, where they regulate downstream signaling components, such as phytochrome interacting factors (PIFs). PIFs are transcription factors that act as repressors of photomorphogenesis; their inhibition by PHYs leads to substantial changes in gene expression. The nuclear function of PHYs, however, has so far been investigated in only a few non-seed plants. Here, we identified putative target genes of PHY signaling in the moss Physcomitrella patens and found light-regulated genes that are putative orthologs of PIF-controlled genes in Arabidopsis. Phylogenetic analyses revealed that an ancestral PIF-like gene was already present in streptophyte algae, i.e., before the water-to-land transition of plants. The PIF homologs in the genome of P. patens resemble Arabidopsis PIFs in their protein domain structure, molecular properties, and physiological effects, albeit with notable differences in the motif-dependent PHY interaction. Our results suggest that P. patens PIFs are involved in PHY signaling. The PHY-PIF signaling node that relays light signals to target genes has been largely conserved during land plant evolution, with evidence of lineage-specific diversification.

BackgroundDoubling the genome contribution of haploid plants has accelerated breeding in most cultivated crop species. Although plant doubled haploids are isogenic in nature, they frequently display unpredictable phenotypes, thus limiting the potential of this technology. Therefore, being able to predict the factors implicated in this phenotypic variability could accelerate the generation of desirable genomic combinations and ultimately plant breeding.ResultsWe use computational analysis to assess the transcriptional and epigenetic dynamics taking place during doubled haploids generation in the genome ofBrassica oleracea. We observe that doubled haploid lines display unexpected levels of transcriptional and epigenetic variation, and that this variation is largely due to imbalanced contribution of parental genomes. We reveal that epigenetic modification of transposon-related sequences during DH breeding contributes to the generation of unpredictable yet heritable transcriptional states. Targeted epigenetic manipulation of these elements using dCas9-hsTET3 confirms their role in transcriptional regulation. We have uncovered a hitherto unknown role for parental genome balance in the transcriptional and epigenetic stability of doubled haploids.ConclusionsThis is the first study that demonstrates the importance of parental genome balance in the transcriptional and epigenetic stability of doubled haploids, thus enabling predictive models to improve doubled haploid-assisted plant breeding.

Plants synthesize and release specialized metabolites into their environment that can serve as chemical cues for other organisms. Metabolites that are released from the roots are important factors in determining which microorganisms will colonize the root and become part of the plant rhizosphere microbiota. Root exudates are often further converted by soil microorganisms, which can result in the formation of toxic compounds. How individual members of the plant rhizosphere respond to individual compounds and how the differential response of individual microorganisms contributes to the response of a microbial community remains unclear. Here, we investigated the impact of derivatives of one class of plant root exudates, benzoxazinoids, which are released by important crops such as wheat and maize, on a collection of 180 root-associated bacteria. We show that phenoxazine, derived from benzoxazinoids, inhibits the growth of root-associated bacteria in vitro in a strain-specific manner, with sensitive and resistant isolates in most of the studied clades. Synthetic bacterial communities that were assembled from only resistant isolates were more resilient to chemical perturbations than communities comprised of only sensitive members. Isolates that were shared between different communities showed stable interactions, independent of the overall community composition. On the other hand, we could attribute differential community development to differences in interactions formed by closely related representatives of the same bacterial genus. Our findings highlight the fact that profiling isolate collections can aid the rational design of synthetic communities. Moreover, our data show that simplified in vitro community systems are able to recapitulate observations on the influence of metabolite exudation on the structure of root-associated communities, thus providing an avenue for reductionist explorations of the rhizosphere biology in defined, host-free settings.

Inducible epigenetic changes in eukaryotes are believed to enable rapid adaptation to environmental fluctuations. We have found distinct regions of the Arabidopsis genome that are susceptible to DNA (de)methylation in response to hyperosmotic stress. The stress-induced epigenetic changes are associated with conditionally heritable adaptive phenotypic stress responses. However, these stress responses are primarily transmitted to the next generation through the female lineage due to widespread DNA glycosylase activity in the male germline, and extensively reset in the absence of stress. Using the CNI1/ATL31 locus as an example, we demonstrate that epigenetically targeted sequences function as distantly-acting control elements of antisense long non-coding RNAs, which in turn regulate targeted gene expression in response to stress. Collectively, our findings reveal that plants use a highly dynamic maternal 'short-term stress memory' with which to respond to adverse external conditions. This transient memory relies on the DNA methylation machinery and associated transcriptional changes to extend the phenotypic plasticity accessible to the immediate offspring.

Summary Immune systems respond to “non-self” molecules termed microbe-associated molecular patterns (MAMPs). Microbial genes encoding MAMPs have adaptive functions and are thus evolutionarily conserved. In the presence of a host, these genes are maladaptive and drive antagonistic pleiotropy (AP) because they promote microbe elimination by activating immune responses. The role AP plays in balancing the functionality of MAMP-coding genes against their immunogenicity is unknown. To address this, we focused on an epitope of flagellin that triggers antibacterial immunity in plants. Flagellin is conserved because it enables motility. Here, we decode the immunogenic and motility profiles of this flagellin epitope and determine the spectrum of amino acid mutations that drives AP. We discover two synthetic mutational tracks that undermine the detection activities of a plant flagellin receptor. These tracks generate epitopes with either antagonist or weaker agonist activities. Finally, we find signatures of these tracks layered atop each other in natural Pseudomonads.

Highlighting an actual plasma copolymerization remains difficult when using infrared or x-ray photoelectron spectroscopies as they provide no evidence of the formation of covalent bonds between the precursors. The combination of high resolution mass spectrometry and mass defect analysis is proposed to characterize plasma (co)polymers of alkyl and perfluorinated acrylates. The mass spectra of the coatings are readily turned into Kendrick mass defect plots unambiguously demonstrating the occurrence of plasma co-oligomers, as suspected from their thermal behavior. Requiring a proper calibration only, the mass defect analysis elegantly transforms an abstruse mass spectrum of plasma polymer in simple point series and constitutes a user-friendly method for data mining.

This article was not made open access when initially published online, which was corrected before print publication. In addition, ORCID links were missing for 12 authors and have been added to the HTML and PDF versions of the article.

Plants differ from animals in their capability to easily regenerate fertile adult individuals from terminally differentiated cells [1]. This unique developmental plasticity is commonly observed in nature where many species can reproduce asexually through the ectopic initiation of organogenic or embryogenic developmental programs [2, 3]. However, it is not currently known if this developmental reprogramming is coupled to a global epigenomic resetting, or what impact it has on the phenotype of the clonal progeny. Here we show that plants asexually propagated via induction of a zygotic developmental program do not fully reset cell-specific epigenetic imprints. These imprints are instead inherited even over multiple rounds of sexual reproduction, becoming fixed in hybrids and resulting in heritable molecular and physiological phenotypes that depend on the founder cell used. Our results demonstrate how novel phenotypic variation in plants can be unlocked through the incomplete reprogramming of cell-specific epigenetic marks during asexual propagation.