Tracing 100 million years of grass genome evolutionary plasticity.

Grasses derive from a family of monocotyledonous plants that includes crops of major economic importance such as wheat, rice, sorghum and barley, sharing a common ancestor some 100 million years ago. The genomic attributes of plant adaptation remain obscure and the consequences of recurrent whole genome duplications (WGD) or polyploidization events, a major force in plant evolution, remain largely speculative. We conducted a comparative analysis of omics data from ten grass species to unveil structural (inversions, fusions, fissions, duplications, substitutions) and regulatory (expression and methylation) basis of genome plasticity, as possible attributes of plant long lasting evolution and adaptation. The present study demonstrates that diverged polyploid lineages sharing a common WGD event often present the same patterns of structural changes and evolutionary dynamics, but these patterns are difficult to generalize across independent WGD events as a result of non-WGD factors such as selection and domestication of crops. Polyploidy is unequivocally linked to the evolutionary success of grasses during the past 100 million years, although it remains difficult to attribute this success to particular genomic consequences of polyploidization, suggesting that polyploids harness the potential of genome duplication, at least partially, in lineage-specific ways. Overall, the present study clearly demonstrates that post-polyploidization reprogramming is more complex than traditionally reported in investigating single species and calls for a critical and comprehensive comparison across independently polyploidized lineages.

A strategy for studying epigenetic diversity in natural populations: proof of concept in poplar and oak.

In the last 20 years, several techniques have been developed for quantifying DNA methylation, the most studied epigenetic marks in eukaryotes, including the gold standard method, whole-genome bisulfite sequencing (WGBS). WGBS quantifies genome-wide DNA methylation but has several inconveniences rendering it less suitable for population-scale epigenetic studies. The high cost of deep sequencing and the large amounts of data generated prompted us to seek an alternative approach. Restricting studies to parts of the genome would be a satisfactory alternative had there not been a major limitation: the need to select upstream targets corresponding to differentially methylated regions as targets. Given the need to study large numbers of samples, we propose a strategy for investigating DNA methylation variation in natural populations, taking into account the structural complexity of genomes, their size, and their content in unique coding regions versus repeated regions as transposable elements. We first identified regions of highly variable DNA methylation in a subset of genotypes representative of the biological diversity in the population by WGBS. We then analysed the variations of DNA methylation in these targeted regions at the population level by sequencing capture bisulfite (SeqCapBis). The entire strategy was then validated by applying it to another species. Our strategy was developed as a proof of concept on natural populations of two forest species: Populus nigra and Quercus petraea.

Low impact of polyploidization on the transcriptome of synthetic allohexaploid wheat.

BACKGROUND: Bread wheat is a recent allohexaploid (genomic constitution AABBDD) that emerged through a hybridization between tetraploid Triticum turgidum (AABB) and diploid Aegilops tauschii (DD) less than 10,000 years ago. The hexaploidization can be re-created artificially, producing synthetic wheat that has been used to study immediate genomic responses to polyploidization. The scale of the consequences of polyploidization, and their mechanism of establishment, remain uncertain. RESULTS: Here we sampled several synthetic wheats from alternative parental genotypes and reciprocal crosses, and examined transcriptomes from two different tissues and successive generations. We did not detect any massive reprogramming in gene expression, with only around 1% of expressed genes showing significant differences compared to their lower-ploidy parents. Most of this differential expression is located on the D subgenome, without consistency in the direction of the expression change. Homoeolog expression bias in synthetic wheat is similar to the pattern observed in the parents. Both differential expression and homoeolog bias are tissue-specific. While up to three families of transposable elements became upregulated in wheat synthetics, their position and distance are not significantly associated with expression changes in proximal genes. DISCUSSION: While only a few genes change their expression pattern after polyploidization, they can be involved in agronomically important pathways. Alternative parental combinations can lead to opposite changes on the same subset of D-located genes, which is relevant for harnessing new diversity in wheat breeding. Tissue specificity of the polyploidization-triggered expression changes indicates the remodelling of transcriptomes in synthetic wheat is plastic and likely caused by regulome interactions rather than permanent changes. We discuss the pitfalls of transcriptomic comparisons across ploidy levels that can inflate the de-regulation signal. CONCLUSIONS: Transcriptomic response to polyploidization in synthetic AABBDD wheat is modest and much lower than some previous estimates. Homoeolog expression bias in wheat allohexaploids is mostly attributed to parental legacy, with polyploidy having a mild balancing effect.

Episodes of gene flow and selection during the evolutionary history of domesticated barley.

BACKGROUND: Barley is one of the founder crops of Neolithic agriculture and is among the most-grown cereals today. The only trait that universally differentiates the cultivated and wild subspecies is 'non-brittleness' of the rachis (the stem of the inflorescence), which facilitates harvesting of the crop. Other phenotypic differences appear to result from facultative or regional selective pressures. The population structure resulting from these regional events has been interpreted as evidence for multiple domestications or a mosaic ancestry involving genetic interaction between multiple wild or proto-domesticated lineages. However, each of the three mutations that confer non-brittleness originated in the western Fertile Crescent, arguing against multiregional origins for the crop. RESULTS: We examined exome data for 310 wild, cultivated and hybrid/feral barley accessions and showed that cultivated barley is structured into six genetically-defined groups that display admixture, resulting at least in part from two or more significant passages of gene flow with distinct wild populations. The six groups are descended from a single founding population that emerged in the western Fertile Crescent. Only a few loci were universally targeted by selection, the identity of these suggesting that changes in seedling emergence and pathogen resistance could represent crucial domestication switches. Subsequent selection operated on a regional basis and strongly contributed to differentiation of the genetic groups. CONCLUSIONS: Identification of genetically-defined groups provides clarity to our understanding of the population history of cultivated barley. Inference of population splits and mixtures together with analysis of selection sweeps indicate descent from a single founding population, which emerged in the western Fertile Crescent. This founding population underwent relatively little genetic selection, those changes that did occur affecting traits involved in seedling emergence and pathogen resistance, indicating that these phenotypes should be considered as 'domestication traits'. During its expansion out of the western Fertile Crescent, the crop underwent regional episodes of gene flow and selection, giving rise to a modern genetic signature that has been interpreted as evidence for multiple domestications, but which we show can be rationalized with a single origin.

Role of genetic introgression during the evolution of cultivated rice (Oryza sativa L.).

BACKGROUND: Models for the origins of cultivated rice currently fall into two groups: ones that identify independent domestications of the indica, japonica and possibly also the aus types, and others that propose that the domestication phenotype was initially acquired by japonica, the underlying alleles then transferred by introgression to other pre-domesticated populations, giving the indica and aus varieties. Identifying the impact of past gene flow on cultivated rice genomes is therefore crucial to distinguishing between these models and understanding the domestication history of rice. To this end, we used population-scale polymorphism data to identify the progenitor gene pools of indica, japonica and aus. Variation shared among the cultivated groups but absent from at least one progenitor population was identified, and genomic blocks putatively transferred by gene flow among cultivated groups mapped. RESULTS: Introgression signals were absent at the major domestication loci (Prog1, Rc, qSH1, qSH3, Sh4) of indica and aus, indicating that these loci were unaffected by gene flow from japonica. Other domestication-related loci (Ghd7, LABA1, Kala4, LG1) show signals of introgression from japonica or indica to aus. There is a strong signal for LABA1 in japonica, possibly indicating introgression from indica. The indica genome is the least affected by gene flow, with just a few short regions with allelic frequencies slightly altered by introgression from japonica. CONCLUSION: Introgression has occurred during the evolution of cultivated rice, but was not responsible for transfer of the key domestication alleles between the cultivated groups. The results are therefore consistent with models in which japonica, indica and aus were domesticated independently, with each of these cultivated groups acquiring the domestication alleles from standing variation in wild rice, without a significant contribution from inter-group gene flow.

A novel mutation conferring the nonbrittle phenotype of cultivated barley.

The nonbrittle rachis, resulting in a seed head which does not shatter at maturity, is one of the key phenotypes that distinguishes domesticated barley from its wild relatives. The phenotype is associated with two loci, Btr1 and Btr2, with all domesticated barleys thought to have either a 1 bp deletion in Btr1 or an 11 bp deletion in Btr2. We used a PCR genotyping method with 380 domesticated barley landraces to identify those with the Btr1 deletion and those with the Btr2 deletion. We discovered two landraces, from Serbia and Greece, that had neither deletion. Instead these landraces possess a novel point mutation in Btr1, changing a leucine to a proline in the protein product. We confirmed that plants carrying this mutation have the nonbrittle phenotype and identified wild haplotypes from the Gaziantep region of southeast Turkey as the closest wild relatives of these two landraces. The presence of a third mutation conferring the nonbrittle phenotype of domesticated barley shows that the origin of this trait is more complex than previously thought, and is consistent with recent models that view the transition to agriculture in southwest Asia as a protracted and multiregional process.

Genetic erosion in domesticated barley and a hypothesis of a North African centre of diversity.

Barley is one of the founder crops of the Neolithic transition in West Asia. While recent advances in genomics have provided a rather detailed picture of barley domestication, there are contradictory views on how the domestication process affected genetic diversity. We set out to revisit this question by integrating public DNA sequencing data from ancient barley and wide collections of extant wild and domesticated accessions. Using two previously overlooked approaches - analyses of chloroplast genomes and genome-wide proportions of private variants - we found that the barley cultivated six millennia ago was genetically unique and more diverse when compared to extant landraces and cultivars. Moreover, the chloroplast genomes revealed a link between the ancient barley, an obscure wild genotype from north-eastern Libya, and a distinct population of barley cultivated in Ethiopia/Eritrea. Based on these results, we hypothesize past existence of a wider North African population that included both wild and cultivated types and suffered from genetic erosion in the past six millennia, likely due to a rapid desertification that ended the Holocene African humid period. Besides providing clues about the origin of Ethiopian landraces, the hypothesis explains the post-domestication loss of diversity observed in barley. Analyses of additional samples will be necessary to resolve the history of African barley and its contribution to the extant cultivated gene pool.

Finding needles in a haystack: identification of inter-specific introgressions in wheat genebank collections using low-coverage sequencing data.

Recently, entire genebank collections of wheat have been extensively characterized with sequencing data. We have identified introgressions using these genotyping-by-sequencing and whole-genome sequencing data. On the basis of our results, we provide information about predicted introgressions at 1-Mb resolution for 9,172 wheat samples as a resource for breeders and scientists. We recommend that all plant genetic resources, including genebank collections, be characterized using a combination of variant calling and introgression prediction. This is necessary to identify potential duplicates in collections efficiently and reliably, and to select promising germplasms with potentially beneficial introgressions for further characterization and prospective breeding application.

Evolutionary history of barley cultivation in Europe revealed by genetic analysis of extant landraces.

BACKGROUND: Understanding the evolution of cultivated barley is important for two reasons. First, the evolutionary relationships between different landraces might provide information on the spread and subsequent development of barley cultivation, including the adaptation of the crop to new environments and its response to human selection. Second, evolutionary information would enable landraces with similar traits but different genetic backgrounds to be identified, providing alternative strategies for the introduction of these traits into modern germplasm. RESULTS: The evolutionary relationships between 651 barley landraces were inferred from the genotypes for 24 microsatellites. The landraces could be divided into nine populations, each with a different geographical distribution. Comparisons with ear row number, caryopsis structure, seasonal growth habit and flowering time revealed a degree of association between population structure and phenotype, and analysis of climate variables indicated that the landraces are adapted, at least to some extent, to their environment. Human selection and/or environmental adaptation may therefore have played a role in the origin and/or maintenance of one or more of the barley landrace populations. There was also evidence that at least some of the population structure derived from geographical partitioning set up during the initial spread of barley cultivation into Europe, or reflected the later introduction of novel varieties. In particular, three closely-related populations were made up almost entirely of plants with the daylength nonresponsive version of the photoperiod response gene PPD-H1, conferring adaptation to the long annual growth season of northern Europe. These three populations probably originated in the eastern Fertile Crescent and entered Europe after the initial spread of agriculture. CONCLUSIONS: The discovery of population structure, combined with knowledge of associated phenotypes and environmental adaptations, enables a rational approach to identification of landraces that might be used as sources of germplasm for breeding programs. The population structure also enables hypotheses concerning the prehistoric spread and development of agriculture to be addressed.

The Chloroplast Land Plant Phylogeny: Analyses Employing Better-Fitting Tree- and Site-Heterogeneous Composition Models.

The colonization of land by descendants of charophyte green algae marked a turning point in Earth history that enabled the development of the diverse terrestrial ecosystems we see today. Early land plants diversified into three gametophyte-dominant lineages, namely the hornworts, liverworts, and mosses, collectively known as bryophytes, and a sporophyte-dominant lineage, the vascular plants, or tracheophytes. In recent decades, the prevailing view of evolutionary relationships among these four lineages has been that the tracheophytes were derived from a bryophyte ancestor. However, recent phylogenetic evidence has suggested that bryophytes are monophyletic, and thus that the first split among land plants gave rise to the lineages that today we recognize as the bryophytes and tracheophytes. We present a phylogenetic analysis of chloroplast protein-coding data that also supports the monophyly of bryophytes. This newly compiled data set consists of 83 chloroplast genes sampled across 30 taxa that include chlorophytes and charophytes, including four members of the Zygnematophyceae, and land plants, that were sampled following a balanced representation of the main bryophyte and tracheophyte lineages. Analyses of non-synonymous site nucleotide data and amino acid translation data result in congruent phylogenetic trees showing the monophyly of bryophytes, with the Zygnematophyceae as the charophyte group most closely related to land plants. Analyses showing that bryophytes and tracheophytes evolved separately from a common terrestrial ancestor have profound implications for the way we understand the evolution of plant life cycles on land and how we interpret the early land plant fossil record.

The mitochondrial phylogeny of land plants shows support for Setaphyta under composition-heterogeneous substitution models.

Congruence among analyses of plant genomic data partitions (nuclear, chloroplast and mitochondrial) is a strong indicator of accuracy in plant molecular phylogenetics. Recent analyses of both nuclear and chloroplast genome data of land plants (embryophytes) have, controversially, been shown to support monophyly of both bryophytes (mosses, liverworts, and hornworts) and tracheophytes (lycopods, ferns, and seed plants), with mosses and liverworts forming the clade Setaphyta. However, relationships inferred from mitochondria are incongruent with these results, and typically indicate paraphyly of bryophytes with liverworts alone resolved as the earliest-branching land plant group. Here, we reconstruct the mitochondrial land plant phylogeny from a newly compiled data set. When among-lineage composition heterogeneity is accounted for in analyses of codon-degenerate nucleotide and amino acid data, the clade Setaphyta is recovered with high support, and hornworts are supported as the earliest-branching lineage of land plants. These new mitochondrial analyses demonstrate partial congruence with current hypotheses based on nuclear and chloroplast genome data, and provide further incentive for revision of how plants arose on land.

Origin of the Aromatic Group of Cultivated Rice (Oryza sativa L.) Traced to the Indian Subcontinent.

The aromatic group of Asian cultivated rice is a distinct population with considerable genetic diversity on the Indian subcontinent and includes the popular Basmati types characterized by pleasant fragrance. Genetic and phenotypic associations with other cultivated groups are ambiguous, obscuring the origin of the aromatic population. From analysis of genome-wide diversity among over 1,000 wild and cultivated rice accessions, we show that aromatic rice originated in the Indian subcontinent from hybridization between a local wild population and examples of domesticated japonica that had spread to the region from their own center of origin in East Asia. Most present-day aromatic accessions have inherited their cytoplasm along with 29-47% of their nuclear genome from the local Indian rice. We infer that the admixture occurred 4,000-2,400 years ago, soon after japonica rice reached the region. We identify aus as the original crop of the Indian subcontinent, indica and japonica as later arrivals, and aromatic a specific product of local agriculture. These results prompt a reappraisal of our understanding of the emergence and development of rice agriculture in the Indian subcontinent.

Origin of rice (Oryza sativa L.) domestication genes.

A number of genes that contribute to the domestication traits of cultivated rice have been identified. These include Sh4, Rc, PROG1 and LABA1, which are associated with non-shattering rachis, white pericarp, erect growth and barbless awns, respectively. The mutations giving rise to the "domestication alleles" of these genes are either invariable in cultivated rice, or have variability that is strictly associated with the phenotypic trait. This observation forms the basis to those current rice domestication models that envisage a single origin for the domesticated phenotype. Such models assume that the domestication alleles are absent or rare in wild rice, emerged under cultivation and spread across all rice groups by introgressive hybridization. We examined whole-genome sequencing datasets for wild and cultivated rice to test the former two assumptions. We found that the rc and laba1 alleles occur in wild rice with broad geographical distribution, and reach frequencies as high as 13 and 15%, respectively. These results are in agreement with previous observations of the prog1 and sh4 domestication alleles in wild populations. We also show that the diversity of the genomic regions surrounding the rc, laba1, prog1 and sh4 alleles in wild accessions is greater than that in cultivated rice, suggesting that these alleles emerged prior to domestication. Our findings indicate that the possibility that independent rice groups obtained identical domestication alleles directly from the wild population needs to be considered.

Three geographically separate domestications of Asian rice.

Domesticated rice (Oryza sativa L.) accompanied the dawn of Asian civilization(1) and has become one of world's staple crops. From archaeological and genetic evidence various contradictory scenarios for the origin of different varieties of cultivated rice have been proposed, the most recent based on a single domestication(2,3). By examining the footprints of selection in the genomes of different cultivated rice types, we show that there were three independent domestications in different parts of Asia. We identify wild populations in southern China and the Yangtze valley as the source of the japonica gene pool, and populations in Indochina and the Brahmaputra valley as the source of the indica gene pool. We reveal a hitherto unrecognized origin for the aus variety in central India or Bangladesh. We also conclude that aromatic rice is a result of a hybridization between japonica and aus, and that the tropical and temperate versions of japonica are later adaptations of one crop. Our conclusions are in accord with archaeological evidence that suggests widespread origins of rice cultivation(1,4). We therefore anticipate that our results will stimulate a more productive collaboration between genetic and archaeological studies of rice domestication, and guide utilization of genetic resources in breeding programmes aimed at crop improvement.

Analyses of charophyte chloroplast genomes help characterize the ancestral chloroplast genome of land plants.

Despite the significance of the relationships between embryophytes and their charophyte algal ancestors in deciphering the origin and evolutionary success of land plants, few chloroplast genomes of the charophyte algae have been reconstructed to date. Here, we present new data for three chloroplast genomes of the freshwater charophytes Klebsormidium flaccidum (Klebsormidiophyceae), Mesotaenium endlicherianum (Zygnematophyceae), and Roya anglica (Zygnematophyceae). The chloroplast genome of Klebsormidium has a quadripartite organization with exceptionally large inverted repeat (IR) regions and, uniquely among streptophytes, has lost the rrn5 and rrn4.5 genes from the ribosomal RNA (rRNA) gene cluster operon. The chloroplast genome of Roya differs from other zygnematophycean chloroplasts, including the newly sequenced Mesotaenium, by having a quadripartite structure that is typical of other streptophytes. On the basis of the improbability of the novel gain of IR regions, we infer that the quadripartite structure has likely been lost independently in at least three zygnematophycean lineages, although the absence of the usual rRNA operonic synteny in the IR regions of Roya may indicate their de novo origin. Significantly, all zygnematophycean chloroplast genomes have undergone substantial genomic rearrangement, which may be the result of ancient retroelement activity evidenced by the presence of integrase-like and reverse transcriptase-like elements in the Roya chloroplast genome. Our results corroborate the close phylogenetic relationship between Zygnematophyceae and land plants and identify 89 protein-coding genes and 22 introns present in the chloroplast genome at the time of the evolutionary transition of plants to land, all of which can be found in the chloroplast genomes of extant charophytes.

Reticulated origin of domesticated emmer wheat supports a dynamic model for the emergence of agriculture in the fertile crescent.

We used supernetworks with datasets of nuclear gene sequences and novel markers detecting retrotransposon insertions in ribosomal DNA loci to reassess the evolutionary relationships among tetraploid wheats. We show that domesticated emmer has a reticulated genetic ancestry, sharing phylogenetic signals with wild populations from all parts of the wild range. The extent of the genetic reticulation cannot be explained by post-domestication gene flow between cultivated emmer and wild plants, and the phylogenetic relationships among tetraploid wheats are incompatible with simple linear descent of the domesticates from a single wild population. A more parsimonious explanation of the data is that domesticated emmer originates from a hybridized population of different wild lineages. The observed diversity and reticulation patterns indicate that wild emmer evolved in the southern Levant, and that the wild emmer populations in south-eastern Turkey and the Zagros Mountains are relatively recent reticulate descendants of a subset of the Levantine wild populations. Based on our results we propose a new model for the emergence of domesticated emmer. During a pre-domestication period, diverse wild populations were collected from a large area west of the Euphrates and cultivated in mixed stands. Within these cultivated stands, hybridization gave rise to lineages displaying reticulated genealogical relationships with their ancestral populations. Gradual movement of early farmers out of the Levant introduced the pre-domesticated reticulated lineages to the northern and eastern parts of the Fertile Crescent, giving rise to the local wild populations but also facilitating fixation of domestication traits. Our model is consistent with the protracted and dispersed transition to agriculture indicated by the archaeobotanical evidence, and also with previous genetic data affiliating domesticated emmer with the wild populations in southeast Turkey. Unlike other protracted models, we assume that humans played an intuitive role throughout the process.