Jon Reinders

Selective epigenetic control of retrotransposition in Arabidopsis

Retrotransposons are mobile genetic elements that populate chromosomes, where the host largely controls their activities. In plants and mammals, retrotransposons are transcriptionally silenced by DNA methylation, which in Arabidopsis is propagated at CG dinucleotides by METHYLTRANSFERASE 1 (MET1). In met1 mutants, however, mobilization of retrotransposons is not observed, despite their transcriptional activation. A post-transcriptional mechanism therefore seems to be preventing retrotransposition. Here we show that a copia-type retrotransposon (Évadé, French for ‘fugitive’) evaded suppression of its movement during inbreeding of hybrid epigenomes consisting of met1- and wild-type-derived chromosomes. Évadé (EVD) reinsertions caused a series of developmental mutations that allowed its identification. Genetic testing of host control of the EVD life cycle showed that transcriptional suppression occurred by CG methylation supported by RNA-directed DNA methylation. On transcriptional reactivation, subsequent steps of the EVD cycle were inhibited by plant-specific RNA polymerase IV/V and the histone methyltransferase KRYPTONITE (KYP). Moreover, genome resequencing demonstrated retrotransposition of EVD but no other potentially active retroelements when this combination of epigenetic mechanisms was compromised. Our results demonstrate that epigenetic control of retrotransposons extends beyond transcriptional suppression and can be individualized for particular elements.

Loss of DNA methylation affects the recombination landscape in Arabidopsis

During sexual reproduction, one-half of the genetic material is deposited in gametes, and a complete set of chromosomes is restored upon fertilization. Reduction of the genetic information before gametogenesis occurs in meiosis, when cross-overs (COs) between homologous chromosomes secure an exchange of their genetic information. COs are not evenly distributed along chromosomes and are suppressed in chromosomal regions encompassing compact, hypermethylated centromeric and pericentromeric DNA. Therefore, it was postulated that DNA hypermethylation is inhibitory to COs. Here, when analyzing meiotic recombination in mutant plants with hypomethylated DNA, we observed unexpected and counterintuitive effects of DNA methylation losses on CO distribution. Recombination was further promoted in the hypomethylated chromosome arms while it was inhibited in heterochromatic regions encompassing pericentromeric DNA. Importantly, the total number of COs was not affected, implying that loss of DNA methylation led to a global redistribution of COs along chromosomes. To determine by which mechanisms altered levels of DNA methylation influence recombination—whether directly in cis or indirectly in trans by changing expression of genes encoding recombination components—we analyzed CO distribution in wild-type lines with randomly scattered and well-mapped hypomethylated chromosomal segments. The results of these experiments, supported by expression profiling data, suggest that DNA methylation affects meiotic recombination in cis. Because DNA methylation exhibits significant variation even within a single species, our results imply that it may influence the evolution of plant genomes through the control of meiotic recombination.

Genome-wide, high-resolution DNA methylation profiling using bisulfite-mediated cytosine conversion

Methylation of cytosines ((m)C) is essential for epigenetic gene regulation in plants and mammals. Aberrant (m)C patterns are associated with heritable developmental abnormalities in plants and with cancer in mammals. We have developed a genome-wide DNA methylation profiling technology employing a novel amplification step for DNA subjected to bisulfite-mediated cytosine conversion. The methylation patterns detected are not only consistent with previous results obtained with (m)C immunoprecipitation (mCIP) techniques, but also demonstrated improved resolution and sensitivity. The technology, named BiMP (for Bisulfite Methylation Profiling), is more cost-effective than mCIP and requires as little as 100 ng of Arabidopsis DNA.

Epigenetic control of transposon transcription and mobility in Arabidopsis

The mobility of genetic elements called transposable elements (TEs) was discovered half a century ago by Barbara McClintock. Although she had recognized them as chromosomal controlling elements, for much of the consequent time TEs were primarily considered as parasites of the host genome. However the recent explosion of discoveries in the fields of genomics and epigenetics have unambiguously shown the importance of TEs in genome function and evolution. Bursts of endogenous TEs have been reported in plants with epigenetic misregulation, revealing the molecular mechanisms underlying their control. We review here the different steps in TE invasion of the host genome involving epigenetic control and environmental stress responses. As TEs propagate in plant genomes and attract epigenetic marks, their neo-insertions can lead to the formation of new, heritable epigenetic variants (epialleles) of genes in their vicinity and impact on host gene regulatory networks. The epigenetic interplay between TE and genes thus plays a crucial role in the TE-host co-evolution.

Heterosis and inbreeding depression of epigenetic Arabidopsis hybrids

We have addressed the possible epigenetic contribution to heterosis using epigenetic inbred lines (epiRILs) with varying levels and distributions of DNA methylation. One line consistently displayed parent-of-origin heterosis for growth-related traits. Genome-wide transcription profiling followed by a candidate gene approach revealed 33 genes with altered regulation in crosses of this line that could contribute to the observed heterosis. Although none of the candidate genes could explain hybrid vigour, we detected intriguing, hybrid-specific transcriptional regulation of the RPP5 gene, encoding a growth suppressor. RPP5 displayed intermediate transcript levels in heterotic hybrids; surprisingly however, with global loss of fitness of their F2 progeny, we observed striking under-representation of the hybrid-like intermediate levels. Thus, in addition to genetic factors contributing to heterosis, our results strongly suggest that epigenetic diversity and epigenetic regulation of transcription play a role in hybrid vigour and inbreeding depression, and also in the absence of parental genetic diversity.

Unlocking the Arabidopsis epigenome.

The patterns of DNA methylation, referred to as the “methylome”, must be faithfully propagated for proper development of plants and mammals. However, it has been unclear to which extent transgenerational epigenetic inheritance will be affected after DNA methylation distribution has been altered. Recently, three reports have addressed this issue in the model plant Arabidopsis thaliana. Here we revisit the results of these experiments addressing the stability of epigenetic inheritance within two populations of epigenetic recombinant inbred lines (epiRILs), in which mosaic epigenomes were subjected to inbreeding for multiple generations. The manner in which the epigenetic variation was induced differed between the two populations, one by adversely affecting chromatin remodeling and the second by impairing the maintenance of DNA methylation, yet the comparison of the results provides a broader view of transgenerational epigenetic inheritance that may find parallels in other organisms.

Parent‐of‐origin control of transgenerational retrotransposon proliferation in Arabidopsis

Retrotransposons are ubiquitous mobile genetic elements constituting a major part of eukaryotic genomes. Yet, monitoring retrotransposition and subsequent copy number increases in multicellular eukaryotes is intrinsically difficult. By following the transgenerational accumulation of a newly activated retrotransposon EVADE (EVD) in Arabidopsis, we noticed fast expansion of activated elements transmitted through the paternal germ line but suppression when EVD‐active copies are maternally inherited. This parent‐of‐origin effect on EVD proliferation was still observed when gametophytes carried mutations for key epigenetic regulators previously shown to restrict EVD mobility. Therefore, the main mechanism preventing active EVD proliferation seems to act through epigenetic control in sporophytic tissues in the mother plant. In consequence, once activated, this retrotransposon proliferates in plant populations owing to suppressed epigenetic control during paternal transmission. This parental gateway might contribute to the occasional bursts of retrotransposon mobilization deduced from the genome sequences of many plant species.

Bisulfite methylation profiling of large genomes

Bisulfite conversion of genomic DNA differentiates cytosines from 5-methylcytosines and, thus, identifies DNA methylation patterns at the single-base level. Here, we review recent developments incorporating high-throughput sequencing of bisulfite-converted DNA for target-specific analyses and genome-wide mapping of plant and mammalian methylomes. These developments include the analysis of human embryonic stem cell and fetal fibroblast methylomes at single-base resolution, which supports the presence of non-CG DNA methylation in wild-type embryonic stem cells and induced pluripotent stem cells. New developments in nanopore sequencing technologies may lead to directly detecting 5-methylcytosine independently of bisulfite conversion, but the current accuracy of this approach remains a limitation. Furthermore, recent investigations detecting 5-hydroxymethylcytosine within mammalian DNA may add yet another level of complexity to the epigenetic code of the methylome.