Native Environment Modulates Leaf Size and Response to Simulated Foliar Shade across Wild Tomato Species

The laminae of leaves optimize photosynthetic rates by serving as a platform for both light capture and gas exchange, while minimizing water losses associated with thermoregulation and transpiration. Many have speculated that plants maximize photosynthetic output and minimize associated costs through leaf size, complexity, and shape, but a unifying theory linking the plethora of observed leaf forms with the environment remains elusive. Additionally, the leaf itself is a plastic structure, responsive to its surroundings, further complicating the relationship. Despite extensive knowledge of the genetic mechanisms underlying angiosperm leaf development, little is known about how phenotypic plasticity and selective pressures converge to create the diversity of leaf shapes and sizes across lineages. Here, we use wild tomato accessions, collected from locales with diverse levels of foliar shade, temperature, and precipitation, as a model to assay the extent of shade avoidance in leaf traits and the degree to which these leaf traits correlate with environmental factors. We find that leaf size is correlated with measures of foliar shade across the wild tomato species sampled and that leaf size and serration correlate in a species-dependent fashion with temperature and precipitation. We use far-red induced changes in leaf length as a proxy measure of the shade avoidance response, and find that shade avoidance in leaves negatively correlates with the level of foliar shade recorded at the point of origin of an accession. The direction and magnitude of these correlations varies across the leaf series, suggesting that heterochronic and/or ontogenic programs are a mechanism by which selective pressures can alter leaf size and form. This study highlights the value of wild tomato accessions for studies of both morphological and light-regulated development of compound leaves, and promises to be useful in the future identification of genes regulating potentially adaptive plastic leaf traits

A Genome-Wide Association Study Identifies Variants Underlying the Arabidopsis thaliana Shade Avoidance Response

Shade avoidance is an ecologically and molecularly well-understood set of plant developmental responses that occur when the ratio of red to far-red light (R∶FR) is reduced as a result of foliar shade. Here, a genome-wide association study (GWAS) in Arabidopsis thaliana was used to identify variants underlying one of these responses: increased hypocotyl elongation. Four hypocotyl phenotypes were included in the study, including height in high R∶FR conditions (simulated sun), height in low R∶FR conditions (simulated shade), and two different indices of the response of height to low R∶FR. GWAS results showed that variation in these traits is controlled by many loci of small to moderate effect. A known PHYC variant contributing to hypocotyl height variation was identified and lists of significantly associated genes were enriched in a priori candidates, suggesting that this GWAS was capable of generating meaningful results. Using metadata such as expression data, GO terms, and other annotation, we were also able to identify variants in candidate de novo genes. Patterns of significance among our four phenotypes allowed us to categorize associations into three groups: those that affected hypocotyl height without influencing shade avoidance, those that affected shade avoidance in a height-dependent fashion, and those that exerted specific control over shade avoidance. This grouping allowed for the development of explicit hypotheses about the genetics underlying shade avoidance variation. Additionally, the response to shade did not exhibit any marked geographic distribution, suggesting that variation in low R∶FR–induced hypocotyl elongation may represent a response to local conditions

Associations with previously-identified SNPs in <i>PHYC</i> and <i>PHYB</i>.

<p>−log10 <i>P</i>-value of Kruskal-Wallis and EMMA associations between hypocotyl height in high R∶FR and candidate SNPs in <i>PHYC</i> and <i>PHYB</i> identified in Balasubramanian et al. <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002589#pgen.1002589-Balasubramanian1" target="_blank">[30]</a> and Filiault et al. <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002589#pgen.1002589-Filiault1" target="_blank">[39]</a>.</p

Associations with <i>a priori</i> candidate genes.

<p>−log10 <i>P</i>-value of most significant Kruskal-Wallis and EMMA associations between hypocotyl phenotypes and <i>a priori</i> candidate genes.</p>*<p>Significance pattern categories: 1 = general control of hypocotyl height, 2 = control of shade avoidance via hypocotyl height, 3 = specific control of shade avoidance response.</p

Parameters from the phenotype mixed effects model.

<p>Estimates of the variance and standard deviation of random effects from the mixed effect model used to generate GWAS phenotypes.</p

Hypocotyl height phenotypes.

<p>(A) Histograms of hypocotyl height for seedings grown under high R∶FR (pink) or low R∶FR (blue) treatments. (B) Hypocotyl height reaction norms of 180 <i>Arabidopsis</i> accessions. Reaction norms for the seven highest-responding accessions (in descending order: 9057, 8242, 6929, 6009, 6914, 6968, 8231) are plotted with red lines, while reaction norms for the seven lowest-responding accession (in ascending order: 6928, 8304, 7515, 6943, 8395, 6916, 8337) are plotted in blue. The three lowest-responding accessions showed a slight negative response to low R∶FR (−0.41, −0.25, and −0.11 millimeters).</p

Manhattan plots of GWAS results.

<p>Genome-wide distribution of the −log10 <i>P</i>-values of SNP/phenotype associations using the Kruskal-Wallis (left panels) and EMMA (right panels) methods. For clarity, only SNPs with a −log10<i>P</i>-value  = 2 are shown. Out of 214548 SNPs assayed, 11102, 6864, 11616, and 5301 SNPs are represented in the Kruskal-Wallis panels (top to bottom) while 2538, 2698, 2399, and 2802 SNPs are represented in the EMMA panels (top to bottom). SNPs are accurately plotted according to their position along the appropriate chromosome. Plotting colors alternate between blue and green in order to facilitate the visualization of individual chromosomes.</p

Relationships between phenotypes.

<p>Correlations between height in high R∶FR and height in low R∶FR (A), height in high R∶FR and response to low R∶FR (B), and height in low R∶FR and response to low R∶FR (C). The kernel density plot in panel D shows the distribution of the corrected response phenotype (residuals from a regression of response to low R∶FR against height in high R∶FR).</p

Correlation between leaf number and environmental variables.

<p><i>r</i> values (Pearson) representing correlation between Leaf Number (LFN) values with environmental variables. Correlation coefficients are given for simulated shade and sun conditions and the ratio of LFN between these treatments (Shade/Sun). Correlation is between either accessions of all species or only accessions of <i>S. arcanum</i> and <i>S. habrochaites</i>. Note the negative correlation between developmental rate and NDVI values under each light treatment and in response to simulated foliar shade. Correlation becomes significant when only <i>S. arcanum</i> and <i>S. habrochaites</i> accessions are considered. *p<0.05, **p<0.01, ***p<0.001.</p