Missing heritability and soft inheritance of morphology and metabolism in Arabidopsis

The plant phenotype is shaped by complex interactions between its genotype and the environment. Although the genotype is stable and determined by the genomic sequence, plants are able to respond flexibly to changes in environmental conditions by orchestrated signal transduction pathways. The genomic sequence may change with each generation through chromosome rearrangements, meiotic recombination and spontaneous mutations. Through natural selection on these randomly induced changes, genotypes become adapted to their local environment. Because different genotypes adapt to different environments, natural variation within species expands in time and gives rise to a wide variety of genotypes and phenotypes. The genetic architecture that specifies the phenotype can be investigated by analyzing different genotypes in the same environment and associate the phenotypic variation with molecular markers that discriminate the genotypes. Recent advances in next-generation sequencing technology enabled the fast sequencing of entire genomes, and in Arabidopsisthalianaalone, more than 1000 different genotypes have been fully resequenced. The sequencing allows the association of phenotypic variation with large numbers of single nucleotide polymorphisms (SNPs) that greatly enhance resolution in genome-wide association studies (GWAS). GWAS on human diseases suffer from missing heritability that is most likely caused by the genetic architecture of the disease traits. Many variants of small effect or rare variants most likely determine a large part of the genetic variation and these variants are difficult to identify in GWAS due to lack of statistical power. In plants, several GWAS have been performed and they have identified previously validated genes and genes involved in monogenic disease resistance, but elucidating quantitative traits such as many agronomic important traits might be problematic in plants as well. Chapter 2 describes a GWA study in which quantitative morphological traits, such as leaf area, flowering time and branching were examined in 350 accessions of Arabidopsis for association with about 200,000 SNPs. The morphological traits showed extensive variation and were highly heritable, but GWA mapping could not identify the genetic variants that explain the heritability. Therefore, missing heritability was addressed using genomic selection models and these models confirmed the quantitative complex architecture of the morphological traits. Based upon these results, the heritability was assumed to be hidden below the significance threshold, and indeed lowering the significance threshold enabled the identification of many candidate genes that have been implicated to play a role in the phenotype directly or indirectly, in previous studies. One candidate gene was studied in more detail; natural variants of ACS11, an ethylene biosynthesis gene, associated significantly with the petiole to leaf length ratio. ACS11is indeed expressed in petioles and ectopically supplied ethylene abolished the difference in the phenotype of natural variants at this locus, strongly suggesting that ACS11is involved in the regulation of petiole growth. However, lowering the significance threshold also increases the number of false-positive associations, non-causal alleles that co-segregate with the trait values. Because regulation of the morphological traits occurs at multiple intermediate levels, increased certainty on the associations can be obtained by performing GWA mapping on the intermediate levels from genotype to phenotype such as gene expression, and protein and metabolite content. Chapter 3 describes a literature survey into the multi-dimensional regulation of metabolic networks that are regulated by inputs from the clock, the communication between cells and between source and sink tissues, and the environment. The metabolic status of the plant can be seen as the final product of the interaction with the environment, and as such, it can serve as a blueprint for growth and development. Chapter 4 describes the abundant variation in enzyme activities and metabolites involved in primary carbon and nitrogen metabolism. The metabolite and enzyme activity data were analyzed together with plant biomass data, and many pleiotropic regulators were identified with opposite effects on primary metabolism and biomass formation. Natural variants in two stress-responsive genes were oppositely associated with biomass and many enzymes and metabolites involved in primary metabolism, suggesting that higher enzyme activities and higher levels of sugars and proteins might be needed to support plant resistance to stress at the expense of growth. Some studies indicated that epigenetic variation, independent of the genetic SNPs, may contribute to missing heritability. Epigenetic inheritance is defined as the inheritance of phenotypic variation to future generations without changes in DNA sequence. Epigenetic variation is caused by variation in chromatin marks such as DNA methylation, histone modifications and small RNAs. Recently, a recombinant inbred line (RIL) population was developed in Arabidopsis where the chromosomes are differentially methylated in lines with an otherwise isogenic background by crossing wild-type Col-0 with a hypomethylated ddm1-2mutant. Chapter 5 describes the epigenetic regulation of morphology and phenotypic plasticity by studying morphological variation in 99 epiRILs under control and saline conditions. The morphology and plasticity trait values were associated with differentially methylated regions (DMRs) that were used as molecular markers in QTL mapping. Many QTLs for various morphological traits and phenotypic plasticity parameters co-located, suggesting pleiotropic epigenetic regulation of growth, morphology and plasticity. Furthermore, methylation variation in the promoter of a salt-tolerance gene, HIGH-AFFINITY K+TRANSPORTER1 (HKT1)associated significantly with leaf area, especially under saline conditions. To gain more insight into the epigenetic regulation of plant growth and morphology, chapter 6 describes the epigenetic regulation of secondary metabolite levels in leaves and flowers and studies the relationship with the morphological traits determined in chapter 5. Many of the QTLs that were found for growth and morphology overlapped with the QTLs for metabolic traits, and suggest pleiotropic regulation. Furthermore, subsets of the metabolites correlated well with the morphological traits and might thus be regulated by the same loci. The majority of metabolite QTLs was detected for glucosinolates and flavonoids in the flowers, and methylation variation was observed for some of the biosynthetic pathway genes of these compounds when comparing Col-0 and ddm1-2, which indicates a role for epigenetic regulation of these biosynthesis pathways. Although stable, natural epialleles have been found in plant species and the environment can induce hypo- and hypermethylation of DNA, it remains elusive whether environmentally-induced epigenetic changes can be inherited to subsequent generations, independent of genetic variation. Chapter 7 describes the transgenerational inheritance of phenotypic variation in progeny derived from a common Arabidopsis founder line. The progeny of stressed parents and grandparents showed variation in morphological traits, metabolite accumulation and gene expression. For example, many salt-responsive genes were up-regulated in progeny of salt-stressed grandparents. The responses to biotic (methyljasmonate) and abiotic (salt) stress differed strongly and this suggests that different environments can cause different transgenerational responses. Because all lines are derived from a single ancestor, epigenetic variation and not DNA variation is most likely causal for the phenotypic variation. Further studies are, however, needed to provide conclusive evidence for transgenerational inheritance. Chapter 8 provides a synthesis of the work and discusses the GWA studies in the light of missing heritability, genetic architecture and the verification of candidate genes. The work on epigenetic regulation of phenotypic plasticity, morphology and metabolism is discussed in relation to Lamarckian soft inheritance that gained new enthusiasm after some recent discoveries in the field of epigenetics. And finally, the metabolomics work is discussed in the light of the growth-defense hypothesis that states that investments in defense occur at the expense of growth.</p

Creación de personaje en la técnica clown de la puesta en escena "Los merolicos"

La creación de personaje en la técnica clown para la puesta en escena “Los merolicos

Multi-dimensional regulation of metabolic networks shaping plant development and performance

The metabolome is an integral part of a plant’s life cycle and determines for a large part its external phenotype. It is the final, internal product of chemical interactions, obtained through developmental, genetic, and environmental inputs, and as such, it defines the state of a plant in terms of development and performance. Understanding its regulation will provide knowledge and new insights into the biochemical pathways and genetic interactions that shape the plant and its surroundings. In this review, we will focus on four dimensions that contribute to the huge diversity of metabolomes and we will illustrate how this diversity shapes the plant in terms of development and performance: (i) temporal regulation: the metabolome is extremely dynamic and temporal changes in the environment can have an immense impact on its composition; (ii) spatial regulation: metabolites can be very specific, in both quantitative and qualitative terms, to specialized organs, tissues, and cell types; (iii) environmental regulation: the metabolic profile of plants is highly dependent on environmental signals, such as light, temperature, and nutrients, and very susceptible to biotic and abiotic stresses; and (iv) genetic regulation: the biosynthesis, structure, and accumulation of metabolites have a genetic origin, and there is quantitative and qualitative variation for metabolomes within a species. We will address the contribution of these dimensions to the wide diversity of metabolomes and highlight how the multi-dimensional regulation of metabolism defines the plant’s phenotyp

Epigenetic variation contributes to environmental adaptation of Arabidopsis thaliana

Epigenetic variation is frequently observed in plants and direct relationships between differences in DNA methylation and phenotypic responses to changing environments have often been described. The identification of contributing genetic loci, however, was until recently hampered by the lack of suitable genome wide mapping resources that specifically segregate for epigenetic marks. The development of epi-RIL populations in the model species Arabidopsis thaliana has alleviated this obstacle, enabling the accurate genetic analysis of epigenetic variation. Comprehensive morphological phenotyping of a ddm1 derived epi-RIL population in different environments and subsequent epi-QTL mapping revealed a high number of epi-QTLs and pleiotropic effects of several DMRs on numerous traits. For a number of these epi-QTLs epistatic interactions could be observed, further adding to the complexity of epigenetic regulation. Moreover, linkage to epigenetic marks indicated a specific role for DNA-methylation variation, rather than TE transposition, in plastic responses to changing environments. These findings provide supportive evidence for a role of epigenetic regulation in evolutionary and adaptive processe

Backcross Populations and Near Isogenic Lines

The development of near isogenic lines (NILs) through repeated backcrossing of genetically distinct parental lines is rather straightforward. Nonetheless, depending on the available resources and the purpose of the lines to be generated, several choices can be made to guide the design of such inbred populations. Here we outline the implications of these choices and provide recommendations for the efficient and proper development of NILs for a number of common scenarios

Arabidopsis semidwarfs evolved from independent mutations in GA20ox1, ortholog to green revolution dwarf alleles in rice and barley.

Semidwarf accessions occur at low frequency across the distribution range of Arabidopsis thaliana and are mainly mutants of the GA5 (GA20ox1) gene, mutations of which originate from wild-type alleles still present in the regions where the mutants were found. We identified the causal mutations by allelism tests and sequencing and performed a detailed population genetics analysis of this variation. Using Fay and Wu¿s H statistics, we obtained indications for local selection of the dwarf alleles. Mutants of functional orthologs of this gene have been selected as the so-called ¿green revolution genes¿ in rice and barley, thus indicating that Arabidopsis natural variation can be a source for the identification of useful genes for plant breeding.Acknowledges support from the Ramón y Cajal Programme of the Ministerio de Ciencia e Innovación (RYC-2011-07847) and the Deutsche Forschungsgemeinschaft Grant SFB680. L.B. was supported by an International Max Planck Research School PhD Fellowship. C.A.-B. was funded by Grant BIO2010-15022 from the Ministerio de Ciencia e Innovación of Spain. R.K. acknowledges funding from the Centre for Biosystems Genomics. Funding was provided by the Max Planck Gesellschaft.Peer Reviewe

Interactions between auxin, microtubules and XTHs mediate green shade-induced petiole elogation in Arabidopsis

Plants are highly attuned to translating environmental changes to appropriate modifications in growth. Such phenotypic plasticity is observed in dense vegetations, where shading by neighboring plants, triggers rapid unidirectional shoot growth (shade avoidance), such as petiole elongation, which is partly under the control of auxin. This growth is fuelled by cellular expansion requiring cell-wall modification by proteins such as xyloglucan endotransglucosylase/hydrolases (XTHs). Cortical microtubules (cMTs) are highly dynamic cytoskeletal structures that are also implicated in growth regulation. The objective of this study was to investigate the tripartite interaction between auxin, cMTs and XTHs in shade avoidance. Our results indicate a role for cMTs to control rapid petiole elongation in Arabidopsis during shade avoidance. Genetic and pharmacological perturbation of cMTs obliterated shade-induced growth and led to a reduction in XTH activity as well. Furthermore, the cMT disruption repressed the shade-induced expression of a specific set of XTHs. These XTHs were also regulated by the hormone auxin, an important regulator of plant developmental plasticity and also of several shade avoidance responses. Accordingly, the effect of cMT disruption on the shade enhanced XTH expression could be rescued by auxin application. Based on the results we hypothesize that cMTs can mediate petiole elongation during shade avoidance by regulating the expression of cell wall modifying proteins via control of auxin distribution

Arabidopsis semidwarfs evolved from independent mutations in GA20ox1, ortholog to green revolution dwarf alleles in rice and barley

Understanding the genetic bases of natural variation for developmental and stress-related traits is a major goal of current plant biology. Variation in plant hormone levels and signaling might underlie such phenotypic variation occurring even within the same species. Here we report the genetic and molecular basis of semidwarf individuals found in natural Arabidopsis thaliana populations. Allelism tests demonstrate that independent loss-offunction mutations at GA locus 5 (GA5), which encodes gibberellin 20-oxidase 1 (GA20ox1) involved in the last steps of gibberellin biosynthesis, are found in different populations from southern, western, and northern Europe; central Asia; and Japan. Sequencing of GA5 identified 21 different loss-of-function alleles causing semidwarfness without any obvious general tradeoff affecting plant performance traits. GA5 shows signatures of purifying selection, whereas GA5 loss-of-function alleles can also exhibit patterns of positive selection in specific populations as shown by Fay and Wu’s H statistics. These results suggest that antagonistic pleiotropy might underlie the occurrence of GA5 loss-of-function mutations in nature. Furthermore, because GA5 is the ortholog of rice SD1 and barley Sdw1/Denso green revolution genes, this study illustrates the occurrence of conserved adaptive evolution between wild A.thaliana and domesticated plant

Epigenetic Basis of Morphological Variation and Phenotypic Plasticity in Arabidopsis thaliana

Epigenetics is receiving growing attention in the plant science community. Epigenetic modifications are thought to play a particularly important role in fluctuating environments. It is hypothesized that epigenetics contributes to plant phenotypic plasticity because epigenetic modifications, in contrast to DNA sequence variation, are more likely to be reversible. The population of decrease in DNA methylation 1-2 (ddm1-2)-derived epigenetic recombinant inbred lines (epiRILs) in Arabidopsis thaliana is well suited for studying this hypothesis, as DNA methylation differences are maximized and DNA sequence variation is minimized. Here, we report on the extensive heritable epigenetic variation in plant growth and morphology in neutral and saline conditions detected among the epiRILs. Plant performance, in terms of branching and leaf area, was both reduced and enhanced by different quantitative trait loci (QTLs) in the ddm1-2 inherited epigenotypes. The variation in plasticity associated significantly with certain genomic regions in which the ddm1-2 inherited epigenotypes caused an increased sensitivity to environmental changes, probably due to impaired genetic regulation in the epiRILs. Many of the QTLs for morphology and plasticity overlapped, suggesting major pleiotropic effects. These findings indicate that epigenetics contributes substantially to variation in plant growth, morphology, and plasticity, especially under stress condition

Data from: Effects of multi-generational stress exposure and offspring environment on the expression and persistence of transgenerational effects in Arabidopsis thaliana

Phenotype data of A. thaliana plants grown in either a field environment or in a climate chamber environment. In the climate chamber environment plants were grown under control conditions or salt stress