Stomata control gas exchanges between the plant and the atmosphere. How natural variation in stomata size and density contributes to resolve trade-offs between carbon uptake and water loss in response to local climatic variation is not yet understood. We developed an automated confocal microscopy approach to characterize natural genetic variation in stomatal patterning in 330 fully sequenced Arabidopsis thaliana accessions collected throughout the European range of the species. We compared this to variation in water-use efficiency, measured as carbon isotope discrimination (δ C). We detect substantial genetic variation for stomata size and density segregating within Arabidopsis thaliana. A positive correlation between stomata size and δ C further suggests that this variation has consequences on water-use efficiency. Genome wide association analyses indicate a complex genetic architecture underlying not only variation in stomatal patterning but also to its covariation with carbon uptake parameters. Yet, we report two novel QTL affecting δ C independently of stomatal patterning. This suggests that, in A. thaliana, both morphological and physiological variants contribute to genetic variance in water-use efficiency. Patterns of regional differentiation and covariation with climatic parameters indicate that natural selection has contributed to shape some of this variation, especially in Southern Sweden, where water availability is more limited in spring relative to summer. These conditions are expected to favour the evolution of drought avoidance mechanisms over drought escape strategies.

A b s t r a c t Stomata control gas exchanges between the plant and the atmosphere. How natural variation in stomata size and density contributes to resolve trade-offs between carbon uptake and water-loss in response to local climatic variation is not yet understood. We developed an automated confocal microscopy approach to characterize natural genetic variation in stomatal patterning in 330 fully-sequenced Arabidopsis thaliana ecotypes collected throughout the European range of the species. We compared this to variation in water-use efficiency, measured as carbon isotope discrimination (δ 13 C). Combined with public genomic and environmental resources, we show that genetic variation for stomata size and density is pervasive in Arabidopsis thaliana. A positive correlation between stomata size and δ 13 C further shows that this variation has consequences on water-use efficiency. Genome-wide association analyses reveal that many loci of small effect contribute not only to variation in stomata patterning but also to its co-variation with carbon uptake parameters. Yet, we report two novel QTL affecting δ 13 C independently of stomata patterning. This suggests that, in A. thaliana, both morphological and physiological variants contribute to genetic variance in water-use efficiency. Patterns of regional differentiation and co-variation with climatic parameters indicate that natural selection has contributed to shape some of this variation, especially in Southern Sweden, where water availability is comparatively more limited in Spring.

Rapid evolution of traits and of plasticity may enable adaptation to climate change, yet solid experimental evidence under natural conditions is scarce. Here, we imposed rainfall manipulations (+30%, control, À30%) for 10 years on entire natural plant communities in two Eastern Mediterranean sites. Additional sites along a natural rainfall gradient and selection analyses in a greenhouse assessed whether potential responses were adaptive. In both sites, our annual target species Biscutella didyma consistently evolved earlier phenology and higher reproductive allocation under drought. Multiple arguments suggest that this response was adaptive: it aligned with theory, corresponding trait shifts along the natural rainfall gradient, and selection analyses under differential watering in the greenhouse. However, another seven candidate traits did not evolve, and there was little support for evolution of plasticity. Our results provide compelling evidence for rapid adaptive evolution under climate change. Yet, several non-evolving traits may indicate potential constraints to full adaptation.

During range expansion, edge populations are expected to face increased genetic drift, which in turn can alter and potentially compromise adaptive dynamics, preventing the removal of deleterious mutations and slowing down adaptation. Here, we contrast populations of the European sub-species Arabidopsis lyrata ssp petraea, which expanded its Northern range after the last glaciation. We document a sharp decline in effective population size in the range-edge population and observe that non-synonymous variants segregate at higher frequencies. We detect a 4.9% excess of derived non-synonymous variants per individual in the range-edge population, suggesting an increase of the genomic burden of deleterious mutations. Inference of the fitness effects of mutations and modeling of allele frequencies under the explicit demographic history of each population predicts a depletion of rare deleterious variants in the range-edge population, but an enrichment for fixed ones, consistent with the bottleneck effect. However, the demographic history of the range-edge population predicts a small net decrease in per-individual fitness. Consistent with this prediction, the range-edge population is not impaired in its growth and survival measured in a common garden experiment. We further observe that the allelic diversity at the self-incompatibility locus, which ensures strict outcrossing and evolves under negative frequency-dependent selection, has remained unchanged. Genomic footprints indicative of selective sweeps are broader in the Northern population but not less frequent. We conclude that the outcrossing species A. lyrata ssp petraea shows a strong resilience to the effect of range expansion.