Daniele L. Filiault

Amino acid polymorphisms in Arabidopsis phytochrome B cause differential responses to light

Plants have a sophisticated system for sensing and responding to their light environment. The light responses of populations and species native to different habitats show adaptive variation; understanding the mechanisms underlying photomorphogenic variation is therefore of significant interest. In Arabidopsis thaliana, phytochrome B (PHYB) is the dominant photoreceptor for red light and plays a major role in white light. Because PHYB has been proposed as a candidate gene for several quantitative trait loci (QTLs) affecting light response, we have investigated sequence and functional variation in Arabidopsis PHYB. We examined PHYB sequences in 33 A. thaliana individuals and in the close relative Arabidopsis lyrata. From 14 nonsynonymous polymorphisms, we chose 5 for further study based on previous QTL studies. In a larger collection of A. thaliana accessions, one of these five polymorphisms, I143L, was associated with variation in red light response. We used transgenic analysis to test this association and confirmed experimentally that natural PHYB polymorphisms cause differential plant responses to light. Furthermore, our results show that allelic variation of PHYB activity is due to amino acid rather than regulatory changes. Together with earlier studies linking variation in light sensitivity to photoreceptor genes, our work suggests that photoreceptors may be a common target of natural selection.

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.

Multiple FLC haplotypes defined by independent cis-regulatory variation underpin life history diversity in Arabidopsis thaliana

Relating molecular variation to phenotypic diversity is a central goal in evolutionary biology. In Arabidopsis thaliana, FLOWERING LOCUS C (FLC) is a major determinant of variation in vernalization--the acceleration of flowering by prolonged cold. Here, through analysis of 1307 A. thaliana accessions, we identify five predominant FLC haplotypes defined by noncoding sequence variation. Genetic and transgenic experiments show that they are functionally distinct, varying in FLC expression level and rate of epigenetic silencing. Allelic heterogeneity at this single locus accounts for a large proportion of natural variation in vernalization that contributes to adaptation of A. thaliana.

Rapid creation of Arabidopsis doubled haploid lines for quantitative trait locus mapping

Quantitative trait loci (QTL) mapping is a powerful tool for investigating the genetic basis of natural variation. QTL can be mapped using a number of different population designs, but recombinant inbred lines (RILs) are among the most effective. Unfortunately, homozygous RIL populations are time consuming to construct, typically requiring at least six generations of selfing starting from a heterozygous F1. Haploid plants produced from an F1 combine the two parental genomes and have only one allele at every locus. Converting these sterile haploids into fertile diploids (termed “doubled haploids,” DHs) produces immortal homozygous lines in only two steps. Here we describe a unique technique for rapidly creating recombinant doubled haploid populations in Arabidopsis thaliana: centromere-mediated genome elimination. We generated a population of 238 doubled haploid lines that combine two parental genomes and genotyped them by reduced representation Illumina sequencing. The recombination rate and parental allele frequencies in our population are similar to those found in existing RIL sets. We phenotyped this population for traits related to flowering time and for petiole length and successfully mapped QTL controlling each trait. Our work demonstrates that doubled haploid populations offer a rapid, easy alternative to RILs for Arabidopsis genetic analysis.

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.

Multiple alleles at a single locus control seed dormancy in Swedish Arabidopsis

Seed dormancy is a complex life history trait that determines the timing of germination and is crucial for local adaptation. Genetic studies of dormancy are challenging, because the trait is highly plastic and strongly influenced by the maternal environment. Using a combination of statistical and experimental approaches, we show that multiple alleles at the previously identified dormancy locus DELAY OF GERMINATION1 jointly explain as much as 57% of the variation observed in Swedish Arabidopsis thaliana, but give rise to spurious associations that seriously mislead genome-wide association studies unless modeled correctly. Field experiments confirm that the major alleles affect germination as well as survival under natural conditions, and demonstrate that locally adaptive traits can sometimes be dissected genetically. DOI: http://dx.doi.org/10.7554/eLife.22502.001

Auxin signaling is a common factor underlying natural variation in tomato shade avoidance

Light is an essential resource for photosynthesis. Limitation of light by shade from plant neighbors can induce a light competition program known as the shade avoidance response (SAR), thereby altering plant growth and development for the sake of survival. Natural genetic variation in SAR is found in plants adapted to distinct environments, including domesticated tomato Solanum lycopersicum and its wild relative Solanum pennellii. QTL mapping was used to examine variation of the SAR between these two species. We found organ specific responses in the elongation of the stem and petiole, including developmental acceleration of growth. Through RNAseq analysis we identified a number of ILs with reduced expression of auxin-related genes in shade treatment. These same ILs display a shade tolerant phenotype in stem growth and overall height. We also identified ILs with altered SAR expression of cell wall expansion genes, although these genotypes had no accompanying alteration in phenotype. Examination of weighted gene co-expression Connectivity networks in sun- and shade-treated plants revealed Connectivity changes in auxin and light signaling genes; this result was supported by the Identification of motifs within the promoters of a subset of shade-responsive genes that were enriched in light signaling, developmental pathways, and auxin responsive transcriptional domains. The Identification ofboth systemic and organ-specific shade tolerance in the ILs, as well as associated changes in the transcriptome, has the potential to inform future studies for breeding plants able to be grown closely (while neighbor-shaded), yet still maintaining high yield. Summary Growth plasticity in response to shade involves expression of specific auxin signaling and cell wall expansion genes, and shade avoidance QTL affect both stem elongation and developmental rate.

The Aquilegia genome: adaptive radiation and an extraordinarily polymorphic chromosome with a unique history

The columbine genus Aquilegia is a classic example of an adaptive radiation, involving a wide variety of pollinators and habitats. Here we present the genome assembly of A. coerulea ‘Goldsmith’, complemented by high-coverage sequencing data from 10 wild species covering the world-wide distribution. Our analyses reveal extensive allele sharing among species and demonstrate that introgression and selection played a role in the Aquilegia radiation. We also present the remarkable discovery that the evolutionary history of an entire chromosome differs from that of the rest of the genome - a phenomenon which we do not fully understand, but which highlights the need to consider chromosomes in an evolutionary context.

Plant genes influence microbial hubs that shape beneficial leaf communities

Although the complex interactions between hosts and microbial associates are increasingly well documented, we still know little about how and why hosts shape microbial communities in nature. We characterized the leaf microbiota within 200 clonal accessions in eight field experiments and detected effects of both local environment and host genotype on community structure. Within environments, hosts’ genetics preferentially associate with a core of ubiquitous microbial hubs that, in turn, structure the community. These microbial hubs correlate with host performance, and a GWAS revealed strong candidate genes for the host factors impacting heritable hubs. Our results reveal how selection may act to enhance fitness through microbial associations and bolster the possibility of enhancing crop performance through these host factors. Text Hosts harbor complex microbial communities that are thought to impact health and development1. This is best studied in human hosts for which the microbiota has been implicated in a variety of diseases including obesity and cancer 2. Efforts are thus underway to determine the host factors shaping these resident populations 3,4 and to use next-generation probiotics to inhibit colonization by pathogens 5. Similarly, in agriculture, there is great hope of shaping the composition of the microbiota in order to mitigate disease and increase crop yield in a sustainable fashion. Indeed, the Food and Agriculture Organization of the United Nations has made the use of biological control and growth promoting microbial associations a clear priority for improving food production 6. Plant associated microbes can be beneficial in many ways including improving access to nutrients, activating or priming the immune system, and competing with pathogens. For example, seeds inoculated with a combination of naturally occurring microbes were recently found to be protected from a sudden-wilt disease that emerged after continuous cropping 7. Thus, it would be advantageous to breed crops that promote the growth of beneficial microbes under a variety of field conditions, a prospect that is made more likely by the demonstration of host genotypic effects on their microbiota 8–10. That said, microbial communities are complex entities that are influenced by the combined impact of host factors, environment and microbe-microbe interactions 11. As a consequence, the extent to which host plants can control microbial communities to their advantage, especially in a natural context, is unclear. Here, we combine large scale field experiments of plant genotypes grown in their natural environments, extensive microbial community analysis, and genome-wide association mapping to (i) disentangle how the influence of the host is distributed among microbial community members, and thus how host variation shapes the microbiota, (ii) propose plant genes and functions that correlate with variation in the microbiota across environmental conditions, and (iii) examine how key microbial associates impact plant fitness. Our motivation is to further the goal of generating plants with an enhanced ability to host beneficial microbial communities.

Verification of Arabidopsis stock collections using SNPmatch, a tool for genotyping high-plexed samples

Large-scale studies such as the Arabidopsis thaliana ‘1,001 Genomes’ Project require routine genotyping of stocks to avoid sample contamination. To genotype samples efficiently and economically, sequencing must be inexpensive and data processing simple. Here we present SNPmatch, a tool that identifies strains (or inbred lines, or accessions) by matching them to a SNP database. We tested the tool by performing low-coverage resequencing of over 2,000 strains from our lab seed stock collection. SNPmatch correctly genotyped samples from 1-fold coverage sequencing data, and could also identify the parents of F1 or F2 individuals. SNPmatch can be run either on the command line or through AraGeno (https://arageno.gmi.oeaw.ac.at), a web interface that permits sample genotyping from a user-uploaded VCF or BED file.