Modeling interactions between transposable elements and the plant epigenetic response: a surprising reliance on element retention

Transposable elements (TEs) compose the majority of angiosperm DNA. Plants counteract TE activity by silencing them epigenetically. One form of epigenetic silencing requires 21-22 nt small interfering RNAs that act to degrade TE mRNA and may also trigger DNA methylation. DNA methylation is reinforced by a second mechanism, the RNA-dependent DNA methylation (RdDM) pathway. RdDM relies on 24 nt small interfering RNAs and ultimately establishes TEs in a quiescent state. These host factors interact at a systems level, but there have been no system level analyses of their interactions. Here, we define a deterministic model that represents the propagation of active TEs, aspects of the host response and the accumulation of silenced TEs. We describe general properties of the model and also fit it to biological data in order to explore two questions. The first is why two overlapping pathways are maintained, given that both are likely energetically expensive. Under our model, RdDM silenced TEs effectively even when the initiation of silencing was weak. This relationship implies that only a small amount of RNAi is needed to initiate TE silencing, but reinforcement by RdDM is necessary to efficiently counter TE propagation. Second, we investigated the reliance of the host response on rates of TE deletion. The model predicted that low levels of deletion lead to few active TEs, suggesting that silencing is most efficient when methylated TEs are retained in the genome, thereby providing one explanation for the large size of plant genomes

Sequencing three crocodilian genomes to illuminate the evolution of archosaurs and amniotes

The International Crocodilian Genomes Working Group (ICGWG) will sequence and assemble the American alligator (Alligator mississippiensis), saltwater crocodile (Crocodylus porosus) and Indian gharial (Gavialis gangeticus) genomes. The status of these projects and our planned analyses are described

The consequences of transposable element and DNA methylation on plant genomes

Plant genomes are not static; they are constantly being transformed by nucleotide substitutions, the propagation of mobile DNA, and epigenetic modifications. In the three chapters of my dissertation, I show how plant genomes are shaped by transposable elements (TEs) and DNA methylation. In the first chapter, I test the hypothesis that DNA plmethylation is involved in differential gene expression between plant tissues. To explore this hypothesis, I measured whole genome DNA methylation and gene expression in leaf and floral bud tissue from Brachypodium distachyon. I found that differential CG methylation in the promoter region explains ~10% of the variation in gene expression between tissues. The second chapter examines the two modes that a plant uses to silence TEs, and I specifically question why both are necessary for efficient TE containment. I address this question by creating a mathematical model of ordinary differential equations that represents the interactions between TE propagation and epigenetic silencing, including DNA methylation. The model suggests that both modes are crucial for efficient silencing, and it also suggests that TE retention leads to more robust silencing. Finally, the third chapter predicts that, because of their deleterious nature, TEs will be ‘purged’ from a lineage that has undergone inbreeding. To test this, I examined the properties of maize genomes that were subjected to inbreeding for six generations. Over a total of 11 inbred lines, I measured genome size with cell flow cytometry and characterized genome content by whole genome sequencing. The results revealed evidence that genome size decline is associated with TE loss in a subset of inbreeding lines and provided an opportunity to consider potential mechanisms for TE removal

The consequences of transposable element and DNA methylation on plant genomes

Plant genomes are not static; they are constantly being transformed by nucleotide substitutions, the propagation of mobile DNA, and epigenetic modifications. In the three chapters of my dissertation, I show how plant genomes are shaped by transposable elements (TEs) and DNA methylation. In the first chapter, I test the hypothesis that DNA plmethylation is involved in differential gene expression between plant tissues. To explore this hypothesis, I measured whole genome DNA methylation and gene expression in leaf and floral bud tissue from Brachypodium distachyon. I found that differential CG methylation in the promoter region explains ~10% of the variation in gene expression between tissues. The second chapter examines the two modes that a plant uses to silence TEs, and I specifically question why both are necessary for efficient TE containment. I address this question by creating a mathematical model of ordinary differential equations that represents the interactions between TE propagation and epigenetic silencing, including DNA methylation. The model suggests that both modes are crucial for efficient silencing, and it also suggests that TE retention leads to more robust silencing. Finally, the third chapter predicts that, because of their deleterious nature, TEs will be ‘purged’ from a lineage that has undergone inbreeding. To test this, I examined the properties of maize genomes that were subjected to inbreeding for six generations. Over a total of 11 inbred lines, I measured genome size with cell flow cytometry and characterized genome content by whole genome sequencing. The results revealed evidence that genome size decline is associated with TE loss in a subset of inbreeding lines and provided an opportunity to consider potential mechanisms for TE removal

CG Methylation Covaries with Differential Gene Expression between Leaf and Floral Bud Tissues of Brachypodium distachyon.

DNA methylation has the potential to influence plant growth and development through its influence on gene expression. To date, however, the evidence from plant systems is mixed as to whether patterns of DNA methylation vary significantly among tissues and, if so, whether these differences affect tissue-specific gene expression. To address these questions, we analyzed both bisulfite sequence (BSseq) and transcriptomic sequence data from three biological replicates of two tissues (leaf and floral bud) from the model grass species Brachypodium distachyon. Our first goal was to determine whether tissues were more differentiated in DNA methylation than explained by variation among biological replicates. Tissues were more differentiated than biological replicates, but the analysis of replicated data revealed high (>50%) false positive rates for the inference of differentially methylated sites (DMSs) and differentially methylated regions (DMRs). Comparing methylation to gene expression, we found that differential CG methylation consistently covaried negatively with gene expression, regardless as to whether methylation was within genes, within their promoters or even within their closest transposable element. The relationship between gene expression and either CHG or CHH methylation was less consistent. In total, CG methylation in promoters explained 9% of the variation in tissue-specific expression across genes, suggesting that CG methylation is a minor but appreciable factor in tissue differentiation

Gene expression with respect to DMRs and their direction.

<p>A) A graph of the distribution of gene expression when a DMR is located within a gene and hypermethylated in the Leaf or Floral Bud, or when there is no DMR in the gene (None). For the 25 genes hypermethylated in leaf, we predicted positive values on the <i>y</i>-axis, signaling higher expression in floral bud, but no bias was detected. For the 19 genes hypermethylated in floral bud, we predicted negative values on the <i>y</i>-axis, signaling higher expression in leaf, but again no bias was detected. B) The same graph of differential expression when the gene contains a DMR in its a promoter region. Again, there are no detectable biases in the direction of gene expression relative to genes that do not contain a DMR in their promoter region. C) A graph of differential gene expression when the TE nearest to a gene has a DMR that is hypermethylated in leaf, flower or no (None) DMR. For all graphs, the box plots represent the median, first, and third quartile. The whiskers represent the minimum and maximum, The numbers above the graph refer to sample size in each category.</p

Methylation patterns within promoters and its relationship to gene expression.

<p>Graphs A,B and C present the level of CG, CHG and CHH methylation, respectively, in terms of distance from the Transcription Start Site. Graphs D, E and F compare methylation contexts, as measured by <i>prop</i> statistics in a log scale, within leaf tissue. Floral bud comparisons are not shown but are visually identical. Panels G, H and I compare differential gene expression [log2fold (FKPM_Flower/FKPM_Leaf)] vs. the difference in <i>prop</i> between floral bud and leaf tissue. The correlation values for G and H are in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0150002#pone.0150002.t003" target="_blank">Table 3</a>.</p

The number of potential methylation sites, DMSs and CMSs in each of three sequence contexts (CG, CHG and CHH) throughout the entire <i>brachypodium</i> genome and also for three features separately (Genes, Promoters and TEs).

<p>The number of potential methylation sites, DMSs and CMSs in each of three sequence contexts (CG, CHG and CHH) throughout the entire <i>brachypodium</i> genome and also for three features separately (Genes, Promoters and TEs).</p

Spearman correlation coefficients between <i>prop</i> values within a tissue.

<p>Spearman correlation coefficients between <i>prop</i> values within a tissue.</p

Spearman correlations between the difference in <i>prop</i> values between tissues and the log2 fold change in gene expression.

<p>Spearman correlations between the difference in <i>prop</i> values between tissues and the log2 fold change in gene expression.</p