The manuscript entitled "Assortment of flowering time and defense alleles in natural Arabidopsis thaliana populations suggests co-evolution between defense and vegetative lifespan strategies" by Glander, He, Schmitz, Witten, Telschow, and de Meaux, addresses an extremely challenging and interesting question in plant biology. The authors seek to understand how plants manage to complete their life cycle whilst coping with pathogens activating genetic defense mechanisms during the vegetative growth period. To this end, the authors make use of various tools, such a model to explore how changes in life history can alter the costs and benefits of host defense systems, readily available data sets from collections of natural accessions including flowering time and genome-wide expression profiles, and experimental data to evaluate the correlation between gene expression and flowering time using the well-known Bur-0 x Col-0 Recombinant Inbred Lines. Overall, the authors are able to provide solid evidence that the higher expression of defense genes and the increase of lifespan duration in A. thaliana have been shaped by natural selection. With no doubt, this work widens our understanding on plant evolution. Below, I go through some issues and concerns with the aim to improve this manuscript.
Given that this manuscript deals with flowering time as a proxy for lifespan, it would be useful to see frequency distributions for flowering time for the different sets of accessions used in this study. I also suggest a panel with the geographical position of accessions (the 138 Swedish accessions from Sasaki et al. 2015, and the 144 accessions from Schmitz et al. 2013) along with the frequency distributions for flowering time for these two accessions plus that for the RILs.
In addition and related to the previous point, it seems that the authors work with accessions that did not require vernalization for flowering. It makes sense as a vernalization treatment prevents to quantify lifespan properly. However, I want to draw the authors' attention to the fact that vernalization is a very common requirement for flowering in natural environments, and thus an important aspect of A. thaliana's biology. I missed some discussion about this important fact just to better frame the conclusions of the work. In my opinion it's not a big deal as vernalization in natural conditions can start quite early after germination. In the case of high altitude populations in the Pyrenees, where spring germination events in late winter or early spring are common and crucial for population persistence, temperatures can be very low during germination and recruitment. We know that plants do not require freezing temperatures but cold ones to get vernalized. Normally, genotypes from high altitude environments require a vernalization requirement for flowering. In the greenhouse, these genotypes can take up to 250 days of vegetative growth for flowering, but in the field they may show short lifespans if they emerge in late winter. This may appear paradoxical, but contrasting lab data with field data is always problematic as the sources of variation are very different. Although this point is partially addressed in this manuscript, in my opinion it requires more attention.
Another important aspect that may also enrich this manuscript is the current knowledge on the response of A. thaliana to biotic and abiotic stresses. Genetic defense mechanisms have been evolving along with the biotic and abiotic selective pressures affecting plants. Two empirical studies by Davila-Olivas et al. on the plant's response to different stresses have recently been published providing valuable insight into this important aspect of A. thaliana's evolutionary history (see New Phytologist, 2017, 213: 838-851, doi: 10.1111/nph.14165; Molecular Ecology, 2017, 26: 2959-2977, doi: 10.1111/mec.14100). These two studies show how life history may explain the susceptibility of A. thaliana to different biotic and abiotic stresses, which overall is quite relevant in the context of this manuscript. I'm quite sure that the authors will find interesting findings to enrich their manuscript.
In general terms, I totally agree with the major take-home messages of this study. However, in some parts of the text the authors refer to the effect of past epidemics on the current defense mechanisms. In other words, they try to build a hypothetical scenario on the causes of what they are studying. I find this a bit confusing because it is really difficult to proof it. In my opinion and based on my observations and field data, A. thaliana strongly adapts to its environment by modifying germination time, flowering time, and the correlation between the two traits. Large-scale climatic variation shapes up life history in A. thaliana and in many other terrestrial organisms. Pathogens and insects are less global but much more local, so my hypothesis is that local populations become adapted to biotic stresses after adjusting life history major events to the abiotic environment. It would be important to pose a more specific hypothesis about how defense mechanisms and life history co-evolved in A. thaliana.
Finally, I ignore whether the authors possess data on seed dormancy or germination behavior for the three sets of accessions used in their study. To date, flowering time variation can no longer be understood without seed dormancy variation. It is widely accepted that the two traits are closely related to each other and that they strongly co-evolve. In a context of developmental niche construction, we know that changes in the timing of life-stage transitions determine the environment experienced subsequently, which in turn affects subsequent phenotypes (see the vast work by K. Donohue on this exciting topic). Thus, it would be very interesting to take seed dormancy into account in this study as the co-evolution between flowering time and defense mechanisms cannot exclude seed dormancy and germination timing. As a matter of fact, we are talking about three players in this evolutionary scenario.