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Using cis-regulatory mutations to highlight polygenic adaptation in natural plant systems

Periodic Reporting for period 4 - AdaptoSCOPE (Using cis-regulatory mutations to highlight polygenic adaptation in natural plant systems)

Reporting period: 2020-03-01 to 2021-02-28

Adaptoscope has demonstrated that the molecular basis of Darwinian adaptation can be discovered if the polygenic basis of adaptation is taken into account, thereby reaching its goal in full.

We used the genome-wide distribution of cis-regulatory variants to discover the molecular pathways that are optimized during adaptation, via accumulation of small effect mutations. We first showed that cis-regulatory variants accumulate in different functions in the diverse Arabidopsis lineages. A first analysis provided a proof-of-concept of the approach in complex multicellular plant lineages and was published in 2016 in Molecular Biology and Evolution. We then used this approach to assess the adaptive evolution of drought stress response genes in the Arabidopsis genus and could show that natural selection had promoted increased stress plasticity in expression in A. lyrata (He et al. 2021, Nature Communications).
We further focused on the two closely related species, A. thaliana and A. lyrata and assembled evidence that these species have adapted locally via polygenic selection. We chose these two species because they both evolved locally adapted population in Northern European latitudes but greatly differ in their mode of development, mating system and ecology. In parallel, we investigated signatures of adaptation in each of these two species. In A. thaliana, we showed that decreased growth rate has most likely contributed to local adaptation to the ecological challenges encountered in Northern Latitudes (Takou et al. Journal of Experimental Botany 2018, Wieters et al. Plos Genetics 2021). In A. lyrata, we demonstrated that adaptive evolution had been maintained despite a severe bottleneck (Takou et al. Molecular Biology and Evolution 2021). Both studies confirmed that polygenic adaptation has played an important role in adaptation to local conditions in both species. We also quantified how much genetic variation in gene expression is available for selection in the outcrossing species A. lyrata. This reavealed that only a small portion of the variance in gene expression is available for natural selection. These results further indicated that, in the outcrossing plant species Arabidopsis lyrata, the regulatory networks controlling plant development were particularly prone to the emergence of phenotypic variation that cannot easily be selected upon, whereas gene expression variation available for selection arises through selective relaxation, which promotes the emergence of variation in the population and ultimately fuels adaptation. In A. thaliana, we also showed that we can annotate genes important for complex trait in the genome by using selected sets of mutants in various environments, providing a functional annotation at the ecological level that is useful for interpreting patterns of cis-regulatory accumulation.
Having confirmed evidence for polygenic adaptation in both plant system, we applied the approach we have pionneered and validated in the first half of the project. We identified variants affecting gene expression cis-regulation in leaves of plants during a crucial phase of their development: exponential vegetative growth. Their distribution in clusters of co-expressed genes was quantified and contrasted across regions. Our analysis shows that the functional distribution of cis-regulatory variants is not random, but enriched in specific molecular functions that differ between regions and lineages. This result was obtained both in A. thaliana and in A. lyrata. This project therefore succeeded in identifying the molecular pathways subjected collectively to natural selection, some of which evolved in parallel in A. thaliana and A. lyrata. This approach has the potential to find broad applications in ecology and agriculture because it points to molecular components that when modified provide sustainable adaptation to growth in novel environments.
The goal of this project was to demonstrate that novel aspects of the molecular basis of Darwinian adaptation can be discovered if the polygenic basis of adaptation is taken into account. Thanks to the genome-wide distribution of cis-regulatory variants, we discovered the molecular pathways that are optimized during adaptation via accumulation of small effect mutations.
The project was completed in 6 Objectives:
Objective 1:
Using a combination of phenotypic analysis, genome-wide association studies and population genetics investigations of signatures of selection, we showed that local adaptation is broadly polygenic, both in A. thaliana and A. lyrata
Objective 2: We developed and tested pipelines to identify allele-specific expression differences pointing to cis-regulatory variation. These pipelines were specifically adjusted to the study of intra- and inter-specific cis-regulatory changes in both species. In addition, we tested and implemented various methods to identify the molecular systems in which cis-regulatory variation accumulates. We generated collections of F1 hybrids resulting from inter-population crosses, within and between species of the Arabidopsis genus. These crossing schemes allowed determining the phylogenetic origin of cis-acting variation. The resulting F1 individuals were grown under common garden conditions and leaf material was harvested at the time of exponential plant growth. We used this material to sequence the transcriptome, applied the pipelines we developped, identified cis-regulatory variants for all expressed genes and determined their phylogenetic origin.
Objective 3: We compared the distribution of cis-regulatory variants of various phylogenetic origin and showed that they accumulate non-randomly in different functions in each local population. This non-random distribution points to the molecular components that have been specifically targeted by natural selection.
Objective 4: We tested whether polygenic selection targeted similar molecular components in different plant species. Comparing results obtained in A. lyrata and A. thaliana, we observe that polygenic adaptation, despite the contribution of myriads of small effect mutations, some of the targeted molecular systems evolve in parallel in different locations.
Objective 5: We implemented various approaches to validate the relevance of the targeted molecular systems for adaptation. This included ecological studies of growth variation, analyses of the transcriptome of A. thaliana mutant affected in plant growth, population genetics tests of selective footprints and genetic analyses of complex trait variation in large interspecific mapping populations.
Objective 6: We assembled a database combining population genetics and functional information across the genomes of A. lyrata and A. thaliana. We used this database to compare the genomic properties that influence the emergence of additive and non-additive variance in gene expression and showed that, in A. lyrata, additive variation is significantly enriched among genes experiencing more relaxed selective constraints. The same analysis has been applied to the analysis of cis-regulatory variation.
The main hypothesis of the project was to test whether the study of cis-regulatory variation could help reveal the molecular footprint of polygenic adaptation. This hypothesis is validated and highlights targets of parallel evolution in two species despite their marked differences in ecology and growth habits.
AdaptoSCOPE studies lineage-specific evolution of cis-regulatory variation.