Periodic Reporting for period 4 - LEAP-EXTREME (Local Edaphic Adaptation in Plants through Leveraging an Extremophile Model)
Okres sprawozdawczy: 2023-04-01 do 2024-09-30
The central objective of this project was to understand plant adaptation to local soil composition using the extremophile metal hyperaccumulator species Arabidopsis halleri as a model. The diploid stoloniferous perennial outcrosser A. halleri diverged from A. thaliana five to ten million years ago. Different from A. thaliana in the sister lineage, and also from several other even more closely related species, A. halleri is a so-called metal hyperaccumulator, of which individuals in natural populations can accumulate extremely high metal concentrations (> 0.3% Zn and > 0.01% Cd) in leaf dry biomass. Additionally, the species A. halleri is unique in having colonized metalliferous soils containing toxic levels of Zn, Cd, Pb, and sometimes of Cu, multiple times. Previous work reported very large extents of trait variation both between populations and within some populations of A. halleri. The molecular basis of such variation remained unidentified at the outset of this project. The project goals were (1) to identify the genetic basis of adaptations to local soil composition, (2) to understand the architecture and plasticity of homeostatic functional networks related to soil composition employing transcriptome data, as well as to examine their within-species divergence, and (3) to unravel genome dynamics in A. halleri that may facilitate local edaphic adaptations in this species.
Towards the first goal (1), we re-sequenced using short reads the genomes of ~1,000 field-collected individuals of A. halleri, for most of which we had also quantified multiple relevant plant and environmental parameters in a field survey (Task 1.1). Based on SNP polymorphisms, we revealed details of population structure and demography, and we completed genome-wide association analyses employing various parameters measured in the field for 835 genotypes (DOI: 10.1111/nph.14219). We also conducted large-scale phenotyping experiments including hundreds of field-collected individuals under standardized environmental conditions in a growth chamber. Furthermore, we mapped QTL for metal hypertolerance and leaf ionomic traits in F2 populations of targeted crosses between phenotypically contrasting accessions of A. halleri upon plant cultivation on appropriately composed artificially metal-contaminated soils (Task 1.2). We identified putative causal loci and polymorphisms within QTL, and also within GWAS peaks, and we addressed and, in part, obtained experimental evidence for their biological roles for a subset of them. Our genetic linkage mapping identified QTL regions contributing to the hyperaccumulation of cadmium, a trait which we observed to be confined to only a subset of phylogenetic lineages of A. halleri. Based on another cross, the approach identified QTL regions underlying locally enhanced metal hypertolerance on an extremely highly metal-contaminated soil.
The generation of a number of haplotype-phased genome assemblies of A. halleri genotypes (for example, https://phytozome-next.jgi.doe.gov/info/Ahalleri_v2_1_0(odnośnik otworzy się w nowym oknie)) which are central for our project enabled us to examine the gene content of the identified QTL and, as a first step, of an exemplary subset GWAS peaks. Thus, we identified candidate genes and polymorphisms, for which we subsequently gained additional evidence using a variety of approaches, including also functional experimental confirmation. These results allow us now to generalize on the genetic architecture of local adaptations in A. halleri and the types of polymorphisms and functional alterations of genes selected for. They also allow us to compare between our QTL mapping and the GWAS approach, allowing conclusions on the origin and evolutionary trajectories of the alleles conferring adaptation.
Towards the second goal (2), we have completed obtained transcriptome data for root and shoot tissues of a large number of genetically diverse A. halleri individuals following cultivation on two contrasting soil types in a growth chamber for RNA extraction and sequencing-based transcriptomics (Task 2.1). For a set of four phenotypically contrasting exemplary accessions of A. halleri and A. thaliana (Col-0), time course experiments of the depletion and excess of single metal ions in hydroponics are completed with all replicates (Task 2.2). Transcriptome data are available any analysed for the subset of excess metal treatments. Towards goal (3) we were able to complete a series of experimental evolution experiments and are preparing for the analysis of plants at the genome level.
Results addressing goal (1) were presented at a number of conferences, and two major manuscripts are at an advanced stage of preparation for publication. Results towards goal (2) were presented at several workshops, and we have begun to work on a manuscript. The results towards goal (3) will require some further work. We are also preparing the genome assemblies generated in this project for their public release. Manuscripts containing small subsets of results for all goals have been published.
The generation of a number of haplotype-phased genome assemblies of A. halleri genotypes (for example, https://phytozome-next.jgi.doe.gov/info/Ahalleri_v2_1_0(odnośnik otworzy się w nowym oknie)) which are central for our project enabled us to examine the gene content of the identified QTL and, as a first step, of an exemplary subset GWAS peaks. Thus, we identified candidate genes and polymorphisms, for which we subsequently gained additional evidence using a variety of approaches, including also functional experimental confirmation. These results allow us now to generalize on the genetic architecture of local adaptations in A. halleri and the types of polymorphisms and functional alterations of genes selected for. They also allow us to compare between our QTL mapping and the GWAS approach, allowing conclusions on the origin and evolutionary trajectories of the alleles conferring adaptation.
Towards the second goal (2), we have completed obtained transcriptome data for root and shoot tissues of a large number of genetically diverse A. halleri individuals following cultivation on two contrasting soil types in a growth chamber for RNA extraction and sequencing-based transcriptomics (Task 2.1). For a set of four phenotypically contrasting exemplary accessions of A. halleri and A. thaliana (Col-0), time course experiments of the depletion and excess of single metal ions in hydroponics are completed with all replicates (Task 2.2). Transcriptome data are available any analysed for the subset of excess metal treatments. Towards goal (3) we were able to complete a series of experimental evolution experiments and are preparing for the analysis of plants at the genome level.
Results addressing goal (1) were presented at a number of conferences, and two major manuscripts are at an advanced stage of preparation for publication. Results towards goal (2) were presented at several workshops, and we have begun to work on a manuscript. The results towards goal (3) will require some further work. We are also preparing the genome assemblies generated in this project for their public release. Manuscripts containing small subsets of results for all goals have been published.
Implementing state-of-the-art genome-enabled and other novel methodologies in this wild and biologically complex species required continuous pioneering developments. Furthermore, our work established and generated results from addressing a plant with a natural history distinct from that of other plants studied commonly in similar approaches, which are usually short-lived and selfing.
We identified the simple genetic basis of the most extreme known case of naturally selected enhanced Cd hypertolerance within A. halleri, We completed multi-trait GWAS employing field trait and environmental data for the species A. halleri and integrated the results, in addition to assembling SNP polymorphisms from ca. 1000 genomes of A. halleri, for future use by the research community. We used these results in our own group to comparatively address metal-related, edaphic and climate-related traits. We generated a number of chromosome-scale haplotype-phased genome assemblies. We identified strongly supported causal genes and genetic polymorphisms for exemplary within-species trait variation, including also Cd hyperaccumulation. Our results reveal the genetic architectures and suggest evolutionary scenarios of the traits focused on in our project. Both our GWAS approach and the QTL mapping in A. halleri highlighted the types of genetic variation constituting the predominant underlying basis of local edaphic adaptations. Interestingly, this differs from earlier findings comparing between A. halleri and A. thaliana. We established the most comprehensive comparative set of plant transcriptome data in relation to metal elements as well as relevant genotypes with contrasting phenotypic properties, as a basis of a thoroughly informative identification of transcriptional networks and their natural variation, based on metal homeostasis.
Beyond the progress beyond the state of the art constituted by our scientific findings, our results are critical for future work addressing micro- and macroevolution, and a wide range of comparative approaches. This project delivered novel fundamental insights into the local adaptation of plants and identified large-effect genetic variants with potential for applications in environmental restoration, biotechnology and crop breeding.
We identified the simple genetic basis of the most extreme known case of naturally selected enhanced Cd hypertolerance within A. halleri, We completed multi-trait GWAS employing field trait and environmental data for the species A. halleri and integrated the results, in addition to assembling SNP polymorphisms from ca. 1000 genomes of A. halleri, for future use by the research community. We used these results in our own group to comparatively address metal-related, edaphic and climate-related traits. We generated a number of chromosome-scale haplotype-phased genome assemblies. We identified strongly supported causal genes and genetic polymorphisms for exemplary within-species trait variation, including also Cd hyperaccumulation. Our results reveal the genetic architectures and suggest evolutionary scenarios of the traits focused on in our project. Both our GWAS approach and the QTL mapping in A. halleri highlighted the types of genetic variation constituting the predominant underlying basis of local edaphic adaptations. Interestingly, this differs from earlier findings comparing between A. halleri and A. thaliana. We established the most comprehensive comparative set of plant transcriptome data in relation to metal elements as well as relevant genotypes with contrasting phenotypic properties, as a basis of a thoroughly informative identification of transcriptional networks and their natural variation, based on metal homeostasis.
Beyond the progress beyond the state of the art constituted by our scientific findings, our results are critical for future work addressing micro- and macroevolution, and a wide range of comparative approaches. This project delivered novel fundamental insights into the local adaptation of plants and identified large-effect genetic variants with potential for applications in environmental restoration, biotechnology and crop breeding.