In forest tree populations, the timing of budbreak (TBB) depends on several factors, both environmental and genetic. The genomic architecture underlying this trait is still not fully characterized. So far, common garden experiments have highlighted a few genomic regions with little heritability, while the whole spectrum of TBB variation observed in the wild still remains unexplored. We performed an in situ genome-wide association study (GWAS) to investigate the genetic and environmental bases of phenotypic variation in budbreak in two wild range-edge populations of the temperate continental tree, European beech (Fagus sylvatica L.) surveyed in two consecutive years. We found distinct significant loci between populations and among years, suggesting that local adaptation has occurred and that genotype-by-environment interactions contribute to the observed variation. The phenotypic variance explained by the significant loci is between 0.2% and 54% for field data and 0% to 59% for modelled data, which suggests that while some traits are polygenic, some others are influenced by a few loci with large effects in the population, which demonstrates the relevance of conducting in situ GWAS in multiple populations. Our findings provide novel insights into the complex genetic architecture of TBB in forest trees. The genetic diversity of budbreak control provides opportunities for selection that could be used through assisted migration to help prepare beech forests to face climate change.

Miniature inverted repeat transposable elements (MITEs) are short, non-autonomous transposable elements that have attracted considerable attention over the years due to their ubiquitous presence and functional roles in plant genomes. A growing body of evidence points to a complex and multifaceted interplay between MITEs and host genomes. This review aims to elucidate the diverse roles of MITEs in shaping plant genome architecture, gene expression and adaptability to environmental stresses through different molecular mechanisms such as accommodation of regulatory sequences, promotion of alternative splicing, generation of epialleles and small RNAs, and mediation of structural variation. These examples highlight the functional importance of MITEs in plant genomes and provide directions for future research.

Shifts in rainfall patterns and increasing temperatures associated with climate change are causing widespread forest decline, especially due to the increase and duration of droughts. Tree species may have to quickly adapt to these changing conditions, and epigenetic modifications are expected to play a key role in regulating rapid acclimation responses. In this study, we measured acclimation physiological responses and methylome responses in mature holm oak trees ( Quercus ilex L.) subjected to 15 years of experimental accrued drought (-29% of rainfall) and their respective controls with ambient rainfall. We hypothesized that: i) oak trees exposed to long-term drought will exhibit different foliar traits due to adaptative phenotypic plasticity to drought, ii) methylation levels will differ between the drought and control trees allowing the identification of drought-induced differentially methylated regions (DMRs), and iii) these DMRs correlate with the differences in foliar traits. Our results confirmed all hypotheses. The methylome analysis revealed 84 drought-related DMRs among trees from different precipitation treatments, of which 17 DMRs were significantly associated with measured phenotypic responses. This study provides evidence of the role of epigenetic regulation for tree acclimation responses in natural populations of holm oak facing increased droughts and identified candidate genes potentially involved in drought adaptation.

Abundant extrachromosomal circular DNA (eccDNA) is associated with transposable element (TE) activity. However, how the eccDNA compartment is controlled by epigenetic regulations and what is its impact on the genome is understudied. Here, using long reads, we sequence both the eccDNA compartment and the genome of Arabidopsis thaliana mutant plants affected in DNA methylation and post-transcriptional gene silencing. We detect a high load of TE-derived eccDNA with truncated and chimeric forms. On the genomic side, on top of truncated and full length TE neo-insertions, we detect complex structural variations (SVs) notably at a disease resistance cluster being a natural hotspot of SV. Finally, we serendipitously identify large tandem duplications in hypomethylated plants, suggesting that SVs could have been overlooked in epigenetic mutants. We propose that a high eccDNA load may alter DNA repair pathways leading to genome instability and the accumulation of SVs, at least in plants.

Horizontal transfer (HT) refers to the exchange of genetic material between divergent species by mechanisms other than reproduction. In recent years, several studies have demonstrated HTs in eukaryotes, particularly in the context of parasitic relationships and in model species. However, very little is known about HT in natural ecosystems, especially those involving non-parasitic wild species, and the nature of the ecological relationships that promote these HTs. In this work, we conducted a pilot study investigating HTs by sequencing the genomes of 17 wild non-model species from a natural ecosystem, the Massane forest, located in southern France. To this end, we developed a new computational pipeline called INTERCHANGE that is able to characterize HTs at the whole genome level without prior annotation and directly in the raw sequencing reads. Using this pipeline, we identified 12 HT events, half of which occurred between lianas and trees. We found that mainly low copy number LTR-retrotransposons from the Copia superfamily were transferred between these wild plant species, especially those of the Ivana and Ale lineages. This study revealed a possible new route for HTs between non-parasitic plants and provides new insights into the genomic characteristics of horizontally transferred DNA in plant genomes.

EDITORIAL article Front. Plant Sci., 08 November 2023Sec. Plant Cell Biology Volume 14 - 2023 | https://doi.org/10.3389/fpls.2023.1260391

The keystone of ribosome biogenesis is the transcription of 45S rDNA. The Arabidopsis thaliana genome contains hundreds of 45S rDNA units; however, they are not all transcribed. Notably, 45S rDNA units contain insertions/deletions revealing the existence of heterogeneous rRNA genes and, likely, heterogeneous ribosomes for rRNAs. In order to obtain an overall picture of 45S rDNA diversity sustaining the synthesis of rRNAs and, subsequently, of ribosomes in natura, we took advantage of 320 new occurrences of Arabidopsis thaliana as a metapopulation named At66, sampled from 0 to 1900 m of altitude in the eastern Pyrenees in France. We found that the 45S rDNA copy number is very dynamic in natura and identified new genotypes for both 5′ and 3′ External Transcribed Spacers (ETS). Interestingly, the highest 5′ETS genotype diversity is found in altitude while the highest 3′ETS genotype diversity is found at sea level. Structural analysis of 45S rDNA also shows conservation in natura of specific 5′ETS and 3′ETS sequences/features required to control rDNA expression and the processing of rRNAs. In conclusion, At66 is a worthwhile natural laboratory, and unraveled 45S rDNA diversity represents an interesting starting material to select subsets for rDNA transcription and alter the rRNA composition of ribosomes both intra- and inter-site.

SUMMARYLong terminal repeat retrotransposons (LTR‐RTs) are mobilized via an RNA intermediate using a ‘copy and paste’ mechanism, and account for the majority of repetitive DNA in plant genomes. As a side effect of mobilization, the formation of LTR‐RT‐derived extrachromosomal circular DNAs (eccDNAs) occurs. Thus, high‐throughput sequencing of eccDNA can be used to identify active LTR‐RTs in plant genomes. Despite the release of a reference genome assembly, carrot LTR‐RTs have not yet been thoroughly characterized. LTR‐RTs are abundant and diverse in the carrot genome. We identified 5976 carrot LTR‐RTs, 2053 and 1660 of which were attributed toCopiaandGypsysuperfamilies, respectively. They were further classified into lineages, families and subfamilies. More diverse LTR‐RT lineages, i.e. lineages comprising many low‐copy‐number subfamilies, were more frequently associated with genic regions. Certain LTR‐RT lineages have been recently active inDaucus carota. In particular, low‐copy‐number LTR‐RT subfamilies, e.g. those belonging to theDcAlelineage, have significantly contributed to carrot genome diversity as a result of continuing activity. We utilized eccDNA sequencing to identify and characterize twoDcAlesubfamilies,Alex1andAlex3, active in carrot callus. We documented 14 and 32de novoinsertions ofAlex1andAlex3, respectively, which were positioned in non‐repetitive regions.

In recent decades, a profound conceptual transformation has occurred comprising different areas of biological research, leading to a novel understanding of life processes as much more dynamic and changeable. Discoveries in plants and animals, as well as novel experimental approaches, have prompted the research community to reconsider established concepts and paradigms. This development was taken as an incentive to organise a workshop in May 2014 at the Academia Nazionale dei Lincei in Rome. There, experts on epigenetics, regeneration, neuroplasticity, and computational biology, using different animal and plant models, presented their insights on important aspects of a dynamic architecture of life, which comprises all organisational levels of the organism. Their work demonstrates that a dynamic nature of life persists during the entire existence of the organism and permits animals and plants not only to fine-tune their response to particular environmental demands during development, but underlies their continuous capacity to do so. Here, a synthesis of the different findings and their relevance for biological thinking is presented. distributed along the central body column (framed box). By contrast cells at the apical and basal extremities are terminally differentiated. A highly dynamic homeostasis with continuous renewal of stem cells supports the fitness and the low senescence of Hydra . Animals survive long periods of starvation as well as the elimination of the nervous system if force-fed. After bisection at any level along the body column, the stump is able to reestablish an organizer centre and regenerate the missing part, either basal or apical. However, some Hydra oligactis strains show a very low level of plasticity: they do not adapt to the loss of neuronal progenitors induced by the transfer to cold temperature, they rapidly lose the ability to regenerate, undergo aging, and finally die in a couple of months. A group of experts met at Accademia dei Lincei to discuss new concepts emerging from experimental data on the dynamic process of living organisms particularly as they relate to differentiation, adaptation, plasticity and memory. The report provides snapshots from selected fields of biology illustrating the paradigm shift that has occurred over the last two decades concerning the dynamics of a variety of molecular and cellular processes. The view that emerges is one of permanent instability that allows for dynamic adaptations to environmental cues. The set of process underlying this instability most likely provides the key to species survival and adaptation, but also, and just as importantly, the emergence of individuality. The paragraphs reporting on the various presentations are sharp and informative and the introductory and concluding ones provide a faithful synthesis of the questions at stake and their elaboration. The only aspect that could have been treated more directly would have been that of synaptic plasticity in the adult which represents undoubtedly an important element of the dynamic architecture of life. This review article represents a remarkable scholarly effort to provide novel insights on a paradigm shift about our understanding of the dynamics of life. Competing Interests: No competing interests were disclosed. I confirm that I have read this submission and believe that I have an appropriate level of expertise to confirm that it is of an acceptable scientific standard. This report summarises the key points of a recent meeting bringing together scientists working on diverse biological systems linked mainly by their approach to understanding the dynamic aspects of biological states. The report highlights the importance of reversibility of differentiation states and cellular responses to external stimuli as well as the importance of this in the context of adaptive evolution. Clearly there are many unanswered questions and the historian’s perspective pointed out that there is much more experimental work required to formulate general principles linking the described systems. Meetings such as these are of great value in allow abstraction of such principles and are to be encouraged. The inclusion of an historian in such meetings is an innovative approach which could be developed further to allow a real in-depth analysis of how ideas evolve over time. Competing Interests: No competing interests were disclosed. We confirm that we have read this submission and believe that we have an appropriate level of expertise to confirm that it is of an acceptable scientific standard.

SummaryThlaspi 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 non‐coding RNAs, pseudogenes and transposable elements, and highlight tissue‐specific expression and methylation patterns. Resequencing of forty wild accessions provided insights into genome‐wide genetic variation, and QTL regions were identified for a seedling colour phenotype. Altogether, these data will serve as a tool for pennycress improvement in general and for translational research across the Brassicaceae.

Throughout the years, most plant genomic studies were focused on nuclear chromosomes. Extrachromosomal circular DNA (eccDNA) has largely been neglected for decades since its discovery in 1965. While initial research showed that eccDNAs can originate from highly repetitive sequences, recent findings show that many regions of the genome can contribute to the eccDNA pool. Currently, the biological functions of eccDNAs, if any, are a mystery but recent studies have indicated that they can be regulated by different genomic loci and contribute to stress response and adaptation. In this review, we outline current relevant technological developments facilitating eccDNA identification and the latest discoveries about eccDNAs in plants. Finally, we explore the probable functions and future research directions that could be undertaken with respect to different eccDNA sources.

Together with local chromatin structure, gene accessibility, and the presence of transcription factors, gene positioning is implicated in gene expression regulation. Although the basic mechanisms are expected to be conserved in eukaryotes, less is known about the role of gene positioning in plant cells, mainly due to the lack of a highly resolutive approach. In this study, we adapted the use of the ANCHOR system to perform real-time single locus detection in planta . ANCHOR is a DNA-labeling tool derived from the chromosome partitioning system found in many bacterial species. We demonstrated its suitability to monitor a single locus in planta and used this approach to track chromatin mobility during cell differentiation in Arabidopsis thaliana root epidermal cells. Finally, we discussed the potential of this approach to investigate the role of gene positioning during transcription and DNA repair in plants.

Extrachromosomal circular DNA (eccDNA) has been observed in different species for decades, and more and more evidence shows that this specific type of DNA molecules may play an important role in rapid adaptation. Therefore, characterizing the full landscape of eccDNA has become critical, and there are several protocols for enriching eccDNAs and performing short-read or long-read sequencing. However, there is currently no available bioinformatic tool to identify eccDNAs from Nanopore reads. More importantly, the current tools based on Illumina short reads lack an efficient standardized pipeline notably to identify eccDNA originating from repeated loci and cannot be applied to very large genomes. Here, we introduce a comprehensive tool to solve both of these two issues.1 Applying ecc_finder to eccDNA-seq data (either mobilome-seq, Circle-Seq and CIDER-seq) from Arabidopsis, human, and wheat (with genome sizes ranging from 120Mb to 17 Gb), we document the improvement of computational time, sensitivity, and accuracy and demonstrate ecc_finder wide applicability and functionality.

Summary Trees are long‐lived organisms that continuously adapt to their environments, a process in which epigenetic mechanisms are likely to play a key role. Via downregulation of the chromatin remodeler DECREASED IN DNA METHYLATION 1 (DDM1) in poplar (Populus tremula × Populus alba) RNAi lines, we examined how DNA methylation coordinates genomic and physiological responses to moderate water deficit. We compared the growth and drought response of two RNAi‐ddm1 lines to wild‐type (WT) trees under well‐watered and water deficit/rewatering conditions, and analyzed their methylomes, transcriptomes, mobilomes and phytohormone contents in the shoot apical meristem. The RNAi‐ddm1 lines were more tolerant to drought‐induced cavitation but did not differ in height or stem diameter growth. About 5000 differentially methylated regions were consistently detected in both RNAi‐ddm1 lines, colocalizing with 910 genes and 89 active transposable elements. Under water deficit conditions, 136 differentially expressed genes were found, including many involved in phytohormone pathways; changes in phytohormone concentrations were also detected. Finally, the combination of hypomethylation and drought led to the mobility of two transposable elements. Our findings suggest major roles for DNA methylation in regulation of genes involved in hormone‐related stress responses, and the maintenance of genome integrity through repression of transposable elements.

Active transposable elements (TEs) generate insertion polymorphisms that can be detected through genome resequencing strategies. However, these techniques may have limitations for organisms with large genomes or for somatic insertions. Here, we present a method that takes advantage of the extrachromosomal circular DNA (eccDNA) forms of actively transposing TEs in order to detect and characterize active TEs in any plant or animal tissue. Mobilome-seq consists in selectively amplifying and sequencing eccDNAs. It relies on linear digestion of genomic DNA followed by rolling circle amplification of circular DNA. Both active DNA transposons and retrotransposons can be identified using this technique.

Among the hundreds of ribosomal RNA (rRNA) gene copies organized as tandem repeats in the nucleolus organizer regions (NORs), only a portion is usually actively expressed in the nucleolus and participate in the ribosome biogenesis process. The role of these extra-copies remains elusive, but previous studies suggested their importance in genome stability and global gene expression. Because the nucleolus is also a platform for nuclear organization, we tested the impact of a decreased amount of rRNA gene copies on the Arabidopsis thaliana 3D genome organization and stability, using an A. thaliana line only containing 20% of rRNA gene copies (20rDNA line). Compared to the wild-type Col-0, the 20rDNA line shows several signs of genomic instability, such as variations in 3D genome organization, spontaneous double-strand breaks accumulation, transcriptomic changes, and higher DNA methylation level. Strikingly, using genomic and microscopic approaches, we identified seven large tandem duplications in direct orientation (TDDOs) ranging from 60 kb to 1.44 Mb. As a consequence, more than 600 genes were duplicated, often associated with an increase in their expression level. Among them, we found several upregulated genes involved in plant-pathogen response, which could explain why the 20rDNA line is hyper-resistant to both bacterial and nematode infections. Finally, we show that the TDDOs create gene fusions and/or truncations and we discuss their potential implications on plant genome evolution.

As fixed and long living organisms subjected to repeated environmental stresses, trees have developed mechanisms such as phenotypic plasticity that help them to cope with fluctuating environmental conditions. Here, we tested the role DNA methylation as a hub of integration, linking plasticity and physiological response to water deficit in the shoot apical meristem of the model tree poplar (Populus). Using a reverse genetic approach, we compared hypomethylated RNAi-ddm1 lines to wild-type trees for drought tolerance. An integrative analysis was realized with phytohormone balance, methylomes, transcriptomes and mobilomes. Hypomethylated lines were more tolerant when subjected to moderate water deficit and were intrinsically more tolerant to drought-induced cavitation. The alteration of the DDM1 machinery induced variation in DNA methylation in a cytosine context dependent manner, both in genes and transposable elements. Hypomethylated lines subjected to water deficit showed altered expression of genes involved in phytohormone pathways, such as salicylic acid and modified hormonal balance. Several transposable elements showed stress- and/or line-specific patterns of reactivation, and we could detect copy number variations for two of them in stressed ddm1 lines. Overall, our data highlight two major roles for DNA methylation in the shoot apical meristem: control of stress response and plasticity through transduction of hormone signaling and maintenance of genome integrity through the control of transposable elements.

Rapid plant genome evolution is crucial to adapt to environmental changes. Chromosomal rearrangements and gene copy number variation (CNV) are two important tools for genome evolution and sources for the creation of new genes. However, their emergence takes many generations. In this study, we show that in Arabidopsis thaliana , a significant loss of ribosomal RNA (rRNA) genes with a past history of a mutation for the chromatin assembly factor 1 (CAF1) complex causes rapid changes in the genome structure. Using long-read sequencing and microscopic approaches, we have identified up to 15 independent large tandem duplications in direct orientation (TDDOs) ranging from 60 kb to 1.44 Mb. Our data suggest that these TDDOs appeared within a few generations, leading to the duplication of hundreds of genes. By subsequently focusing on a line only containing 20% of rRNA gene copies (20rDNA line), we investigated the impact of TDDOs on 3D genome organization, gene expression, and cytosine methylation. We found that duplicated genes often accumulate more transcripts. Among them, several are involved in plant–pathogen response, which could explain why the 20rDNA line is hyper-resistant to both bacterial and nematode infections. Finally, we show that the TDDOs create gene fusions and/or truncations and discuss their potential implications for the evolution of plant genomes.

Copia/Ale is the youngest lineage in both Solanum tuberosum and S. commersonii. Within it, we identified nightshade, a new LTR element active in the cultivated potato. From an evolutionary perspective, long-terminal repeat retrotransposons (LTR-RT) activity during stress may be viewed as a mean by which organisms can keep up rates of genetic adaptation to changing conditions. Potato is one of the most important crop consumed worldwide, but studies on LTR-RT characterization are still lacking. Here, we assessed the abundance, insertion time and activity of LTR-RTs in both cultivated Solanum tuberosum and its cold-tolerant wild relative S. commersonii genomes. Gypsy elements were more abundant than Copia ones, suggesting that the former was somehow more successful in colonizing potato genomes. However, Copia elements, and in particular, the Ale lineage, are younger than Gypsy ones, since their insertion time was in average ~ 2 Mya. Due to the ability of LTR-RTs to be circularized by the host DNA repair mechanisms, we identified via mobilome-seq a Copia/Ale element (called nightshade, informal name used for potato family) active in S. tuberosum genome. Our analyses represent a valuable resource for comparative genomics within the Solanaceae, transposon-tagging and for the design of cultivar-specific molecular markers in potato.