Transposon dynamics in the emerging oilseed crop Thlaspi arvense.

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 might contribute to the broad geographic success and adaptive capacity of the emerging oil crop Thlaspi arvense (field pennycress). We classified the TE inventory based on a high-quality genome assembly, estimated the age of retrotransposon TE families and comprehensively assessed their mobilization potential. A survey of 280 accessions from 12 regions across the Northern hemisphere allowed us to quantify over 90,000 TE insertion polymorphisms (TIPs). Their distribution mirrored the 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. Ty3/Athila elements are the main drivers of TE diversity in T. arvense populations, while a single Ty1/Alesia lineage might be particularly important for transcriptome divergence. The number of retrotransposon TIPs is associated with variation at genes related to epigenetic regulation, including an apparent knockout mutation in BROMODOMAIN AND ATPase DOMAIN-CONTAINING PROTEIN 1 (BRAT1), while DNA transposons are associated with variation at the HSP19 heat shock protein gene. 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 of T. arvense.

Sensitive inference of alignment-safe intervals from biodiverse protein sequence clusters using EMERALD.

Sequence alignments are the foundations of life science research, but most innovation so far focuses on optimal alignments, while information derived from suboptimal solutions is ignored. We argue that one optimal alignment per pairwise sequence comparison is a reasonable approximation when dealing with very similar sequences but is insufficient when exploring the biodiversity of the protein universe at tree-of-life scale. To overcome this limitation, we introduce pairwise alignment-safety to uncover the amino acid positions robustly shared across all suboptimal solutions. We implement EMERALD, a software library for alignment-safety inference, and apply it to 400k sequences from the SwissProt database.

Uncovering gene-family founder events during major evolutionary transitions in animals, plants and fungi using GenEra.

We present GenEra ( https://github.com/josuebarrera/GenEra ), a DIAMOND-fueled gene-family founder inference framework that addresses previously raised limitations and biases in genomic phylostratigraphy, such as homology detection failure. GenEra also reduces computational time from several months to a few days for any genome of interest. We analyze the emergence of taxonomically restricted gene families during major evolutionary transitions in plants, animals, and fungi. Our results indicate that the impact of homology detection failure on inferred patterns of gene emergence is lineage-dependent, suggesting that plants are more prone to evolve novelty through the emergence of new genes compared to animals and fungi.

Canalization of genome-wide transcriptional activity in Arabidopsis thaliana accessions by MET1-dependent CG methylation.

BACKGROUND: Despite its conserved role on gene expression and transposable element (TE) silencing, genome-wide CG methylation differs substantially between wild Arabidopsis thaliana accessions. RESULTS: To test our hypothesis that global reduction of CG methylation would reduce epigenomic, transcriptomic, and phenotypic diversity in A. thaliana accessions, we knock out MET1, which is required for CG methylation, in 18 early-flowering accessions. Homozygous met1 mutants in all accessions suffer from common developmental defects such as dwarfism and delayed flowering, in addition to accession-specific abnormalities in rosette leaf architecture, silique morphology, and fertility. Integrated analysis of genome-wide methylation, chromatin accessibility, and transcriptomes confirms that MET1 inactivation greatly reduces CG methylation and alters chromatin accessibility at thousands of loci. While the effects on TE activation are similarly drastic in all accessions, the quantitative effects on non-TE genes vary greatly. The global expression profiles of accessions become considerably more divergent from each other after genome-wide removal of CG methylation, although a few genes with diverse expression profiles across wild-type accessions tend to become more similar in mutants. Most differentially expressed genes do not exhibit altered chromatin accessibility or CG methylation in cis, suggesting that absence of MET1 can have profound indirect effects on gene expression and that these effects vary substantially between accessions. CONCLUSIONS: Systematic analysis of MET1 requirement in different A. thaliana accessions reveals a dual role for CG methylation: for many genes, CG methylation appears to canalize expression levels, with methylation masking regulatory divergence. However, for a smaller subset of genes, CG methylation increases expression diversity beyond genetically encoded differences.

Grafting vigour is associated with DNA de-methylation in eggplant.

In horticulture, grafting is a popular technique used to combine positive traits from two different plants. This is achieved by joining the plant top part (scion) onto a rootstock which contains the stem and roots. Rootstocks can provide resistance to stress and increase plant production, but despite their wide use, the biological mechanisms driving rootstock-induced alterations of the scion phenotype remain largely unknown. Given that epigenetics plays a relevant role during distance signalling in plants, we studied the genome-wide DNA methylation changes induced in eggplant (Solanum melongena) scion using two interspecific rootstocks to increase vigour. We found that vigour was associated with a change in scion gene expression and a genome-wide hypomethylation in the CHH context. Interestingly, this hypomethylation correlated with the downregulation of younger and potentially more active long terminal repeat retrotransposable elements (LTR-TEs), suggesting that graft-induced epigenetic modifications are associated with both physiological and molecular phenotypes in grafted plants. Our results indicate that the enhanced vigour induced by heterografting in eggplant is associated with epigenetic modifications, as also observed in some heterotic hybrids.

noisyR: enhancing biological signal in sequencing datasets by characterizing random technical noise.

High-throughput sequencing enables an unprecedented resolution in transcript quantification, at the cost of magnifying the impact of technical noise. The consistent reduction of random background noise to capture functionally meaningful biological signals is still challenging. Intrinsic sequencing variability introducing low-level expression variations can obscure patterns in downstream analyses. We introduce noisyR, a comprehensive noise filter to assess the variation in signal distribution and achieve an optimal information-consistency across replicates and samples; this selection also facilitates meaningful pattern recognition outside the background-noise range. noisyR is applicable to count matrices and sequencing data; it outputs sample-specific signal/noise thresholds and filtered expression matrices. We exemplify the effects of minimizing technical noise on several datasets, across various sequencing assays: coding, non-coding RNAs and interactions, at bulk and single-cell level. An immediate consequence of filtering out noise is the convergence of predictions (differential-expression calls, enrichment analyses and inference of gene regulatory networks) across different approaches.

Sensitive protein alignments at tree-of-life scale using DIAMOND.

We are at the beginning of a genomic revolution in which all known species are planned to be sequenced. Accessing such data for comparative analyses is crucial in this new age of data-driven biology. Here, we introduce an improved version of DIAMOND that greatly exceeds previous search performances and harnesses supercomputing to perform tree-of-life scale protein alignments in hours, while matching the sensitivity of the gold standard BLASTP.

A Predictive Approach to Infer the Activity and Natural Variation of Retrotransposon Families in Plants.

Plant genomes harbor a particularly rich landscape of repetitive sequences. Transposable elements (TEs) represent a major fraction of this diversity and are intimately linked with plasticity and evolution of genomes across the tree of life (Fedoroff, Science 338:758-767, 2012). Amplification of Long Terminal Repeats (LTR) retrotransposons have shaped the genomic landscape by reshuffling genomic regions, altering gene expression, and providing new regulatory sequences, some of which have been instrumental for crop domestication and breeding (Lisch, Nat Rev Genet 14:49-61, 2013; Vitte et al., Brief Funct Genomics 13:276-295, 2014). While many retrotransposon families are still active within plant genomes, the repetitive nature of retrotransposons has hindered accurate annotation and kingdom-wide predictive assessment of their activity and molecular evolution. While it is natural for the first approach towards a genome annotation to characterize all regions of the genome and associate them with known structures such as particular genes, transposable elements, or other types of non-coding regions, such efforts can result in a large proportion of false-positive annotations when seeking for active loci. To overcome this issue, the next round of annotation efforts needs to include functional annotations based on rigorously defined sequence structures and protein domain compositions. In the context of retrotransposons, such a functional annotation can enable efforts to mobilize particular retrotransposon families in species living today and harness their mutagenic potency for crop improvement (Paszkowski, Curr Opin Biotechnol 32:200-206, 2015). For this purpose, we present a predictive analytical approach to infer the activity and natural variation of retrotransposon families in plants. This is achieved by applying a combination of software and molecular biology tools we developed for functional annotation, activity monitoring, and the assessment of the population structure of particular retrotransposon families in multiple plant species.

How Stress Facilitates Phenotypic Innovation Through Epigenetic Diversity.

Climate adaptation through phenotypic innovation will become the main challenge for plants during global warming. Plants exhibit a plethora of mechanisms to achieve environmental and developmental plasticity by inducing dynamic alterations of gene regulation and by maximizing natural variation through large population sizes. While successful over long evolutionary time scales, most of these mechanisms lack the short-term adaptive responsiveness that global warming will require. Here, we review our current understanding of the epigenetic regulation of plant genomes, with a focus on stress-response mechanisms and transgenerational inheritance. Field and laboratory-scale experiments on plants exposed to stress have revealed a multitude of temporally controlled, mechanistic strategies integrating both genetic and epigenetic changes on the genome level. We analyze inter- and intra-species population diversity to discuss how methylome differences and transposon activation can be harnessed for short-term adaptive efforts to shape co-evolving traits in response to qualitatively new climate conditions and environmental stress.

The Earth BioGenome project: opportunities and challenges for plant genomics and conservation.

Sequencing them all. That is the ambitious goal of the recently launched Earth BioGenome project (Proceedings of the National Academy of Sciences of the United States of America, 115, 4325-4333), which aims to produce reference genomes for all eukaryotic species within the next decade. In this perspective, we discuss the opportunities of this project with a plant focus, but highlight also potential limitations. This includes the question of how to best capture all plant diversity, as the green taxon is one of the most complex clades in the tree of life, with over 300 000 species. For this, we highlight four key points: (i) the unique biological insights that could be gained from studying plants, (ii) their apparent underrepresentation in sequencing efforts given the number of threatened species, (iii) the necessity of phylogenomic methods that are aware of differences in genome complexity and quality, and (iv) the accounting for within-species genetic diversity and the historical aspect of conservation genetics.

Becoming a Selfish Clan: Recombination Associated to Reverse-Transcription in LTR Retrotransposons.

Transposable elements (TEs) are parasitic DNA bits capable of mobilization and mutagenesis, typically suppressed by host's epigenetic silencing. Since the selfish DNA concept, it is appreciated that genomes are also molded by arms-races against natural TE inhabitants. However, our understanding of evolutionary processes shaping TEs adaptive populations is scarce. Here, we review the events of recombination associated to reverse-transcription in LTR retrotransposons, a process shuffling their genetic variants during replicative mobilization. Current evidence may suggest that recombinogenic retrotransposons could beneficially exploit host suppression, where clan behavior facilitates their speciation and diversification. Novel refinements to retrotransposons life-cycle and evolution models thus emerge.

Environmental and epigenetic regulation of Rider retrotransposons in tomato.

Transposable elements in crop plants are the powerful drivers of phenotypic variation that has been selected during domestication and breeding programs. In tomato, transpositions of the LTR (long terminal repeat) retrotransposon family Rider have contributed to various phenotypes of agronomical interest, such as fruit shape and colour. However, the mechanisms regulating Rider activity are largely unknown. We have developed a bioinformatics pipeline for the functional annotation of retrotransposons containing LTRs and defined all full-length Rider elements in the tomato genome. Subsequently, we showed that accumulation of Rider transcripts and transposition intermediates in the form of extrachromosomal DNA is triggered by drought stress and relies on abscisic acid signalling. We provide evidence that residual activity of Rider is controlled by epigenetic mechanisms involving siRNAs and the RNA-dependent DNA methylation pathway. Finally, we demonstrate the broad distribution of Rider-like elements in other plant species, including crops. Our work identifies Rider as an environment-responsive element and a potential source of genetic and epigenetic variation in plants.

Sensitive detection of pre-integration intermediates of long terminal repeat retrotransposons in crop plants.

Retrotransposons have played an important role in the evolution of host genomes1,2. Their impact is mainly deduced from the composition of DNA sequences that have been fixed over evolutionary time2. Such studies provide important 'snapshots' reflecting the historical activities of transposons but do not predict current transposition potential. We previously reported sequence-independent retrotransposon trapping (SIRT) as a method that, by identification of extrachromosomal linear DNA (eclDNA), revealed the presence of active long terminal repeat (LTR) retrotransposons in Arabidopsis3. However, SIRT cannot be applied to large and transposon-rich genomes, as found in crop plants. We have developed an alternative approach named ALE-seq (amplification of LTR of eclDNAs followed by sequencing) for such situations. ALE-seq reveals sequences of 5' LTRs of eclDNAs after two-step amplification: in vitro transcription and subsequent reverse transcription. Using ALE-seq in rice, we detected eclDNAs for a novel Copia family LTR retrotransposon, Go-on, which is activated by heat stress. Sequencing of rice accessions revealed that Go-on has preferentially accumulated in Oryza sativa ssp. indica rice grown at higher temperatures. Furthermore, ALE-seq applied to tomato fruits identified a developmentally regulated Gypsy family of retrotransposons. A bioinformatic pipeline adapted for ALE-seq data analyses is used for the direct and reference-free annotation of new, active retroelements. This pipeline allows assessment of LTR retrotransposon activities in organisms for which genomic sequences and/or reference genomes are either unavailable or of low quality.

SecretSanta: flexible pipelines for functional secretome prediction.

Motivation: The secretome denotes the collection of secreted proteins exported outside of the cell. The functional roles of secreted proteins include the maintenance and remodelling of the extracellular matrix as well as signalling between host and non-host cells. These features make secretomes rich reservoirs of biomarkers for disease classification and host-pathogen interaction studies. Common biomarkers are extracellular proteins secreted via classical pathways that can be predicted from sequence by annotating the presence or absence of N-terminal signal peptides. Several heterogeneous command line tools and web-interfaces exist to identify individual motifs, signal sequences and domains that are either characteristic or strictly excluded from secreted proteins. However, a single flexible secretome-prediction workflow that combines all analytic steps is still missing. Results: To bridge this gap the SecretSanta package implements wrapper and parser functions around established command line tools for the integrative prediction of extracellular proteins that are secreted via classical pathways. The modularity of SecretSanta enables users to create tailored pipelines and apply them across the whole tree of life to facilitate comparison of secretomes across multiple species or under various conditions. Availability and implementation: SecretSanta is implemented in the R programming language and is released under GPL-3 license. All functions have been optimized and parallelized to allow large-scale processing of sequences. The open-source code, installation instructions and vignette with use case scenarios can be downloaded from https://github.com/gogleva/SecretSanta. Supplementary information: Supplementary data are available at Bioinformatics online.

myTAI: evolutionary transcriptomics with R.

Motivation: Next Generation Sequencing (NGS) technologies generate a large amount of high quality transcriptome datasets enabling the investigation of molecular processes on a genomic and metagenomic scale. These transcriptomics studies aim to quantify and compare the molecular phenotypes of the biological processes at hand. Despite the vast increase of available transcriptome datasets, little is known about the evolutionary conservation of those characterized transcriptomes. Results: The myTAI package implements exploratory analysis functions to infer transcriptome conservation patterns in any transcriptome dataset. Comprehensive documentation of myTAI functions and tutorial vignettes provide step-by-step instructions on how to use the package in an exploratory and computationally reproducible manner. Availability and implementation: The open source myTAI package is available at https://github.com/HajkD/myTAI and https://cran.r-project.org/web/packages/myTAI/index.html. Contact: hgd23@cam.ac.uk. Supplementary information: Supplementary data are available at Bioinformatics online.

High-frequency recombination between members of an LTR retrotransposon family during transposition bursts.

Retrotransposons containing long terminal repeats (LTRs) form a substantial fraction of eukaryotic genomes. The timing of past transposition can be estimated by quantifying the accumulation of mutations in initially identical LTRs. This way, retrotransposons are divided into young, potentially mobile elements, and old that moved thousands or even millions of years ago. Both types are found within a single retrotransposon family and it is assumed that the old members will remain immobile and degenerate further. Here, we provide evidence in Arabidopsis that old members enter into replication/transposition cycles through high rates of intra-family recombination. The recombination occurs pairwise, resembling the formation of recombinant retroviruses. Thus, each transposition burst generates a novel progeny population of chromosomally integrated LTR retrotransposons consisting of pairwise recombination products produced in a process comparable the sexual exchange of genetic information. Our observations provide an explanation for the reported high rates of sequence diversification in retrotransposons.

Developmental Restriction of Retrotransposition Activated in Arabidopsis by Environmental Stress.

Retrotransposons (RTs) can rapidly increase in copy number due to periodic bursts of transposition. Such bursts are mutagenic and thus potentially deleterious. However, certain transposition-induced gain-of-function or regulatory mutations may be of selective advantage. How an optimal balance between these opposing effects arises is not well characterized. Here, we studied transposition bursts of a heat-activated retrotransposon family in Arabidopsis We recorded a high inter and intraplant variation in the number and chromosomal position of new insertions, which usually did not affect plant fertility and were equally well transmitted through male and female gametes, even though 90% of them were within active genes. We found that a highly heterogeneous distribution of these new retroelement copies result from a combination of two mechanisms, of which the first prevents multiple transposition bursts in a given somatic cell lineage that later contributes to differentiation of gametes, and the second restricts the regulatory influence of new insertions toward neighboring chromosomal DNA. As a whole, such regulatory characteristics of this family of RTs ensure its rapid but stepwise accumulation in plant populations experiencing transposition bursts accompanied by high diversity of chromosomal sites harboring new RT insertions.

Cross-kingdom comparison of the developmental hourglass.

The developmental hourglass model has its foundations in classic anatomical studies by von Baer and Haeckel. In this context, even the conservation of animal body plans has been explained by evolutionary constraints acting on mid-embryogenic development. Recent studies have shown that developmental hourglass patterns also exist on the transcriptomic level, mirroring the corresponding morphological patterns. The identification of similar patterns in embryonic, post-embryonic, and life cycle spanning transcriptomes in plant and fungus development, however, contradict the notion of a direct coupling between morphological and molecular patterns. To explain the existence of hourglass patterns across kingdoms and developmental processes, we propose the organizational checkpoint model that integrates the developmental hourglass model into a framework of transcriptome switches.