Abstract:Retrotransposons are mobile genetic elements abundant in plant and animal genomes. While efficiently silenced by the epigenetic machinery, they can be reactivated upon stress or during development. Their level of transcription not reflecting their transposition ability, it is thus difficult to evaluate their contribution to the active mobilome. Here we applied a simple methodology based on the high throughput sequencing of extrachromosomal circular DNA (eccDNA) forms of active retrotransposons to characterize … Show more
“…eccDNA is a byproduct of the LTR retrotransposon life cycle [20]. Using mobilome sequencing, which comprises a specific amplification step of circular DNA followed by high-throughput sequencing to identify eccDNA derived from active LTR retrotransposons [2], we found that only ONSEN was activated by HS in combination with A&Z (Additional file 1: Figure S2). Confirming our qPCR data, more ONSEN -specific reads were detected in the presence of A and Z in the medium.Fig.…”
BackgroundRetrotransposons play a central role in plant evolution and could be a powerful endogenous source of genetic and epigenetic variability for crop breeding. To ensure genome integrity several silencing mechanisms have evolved to repress retrotransposon mobility. Even though retrotransposons fully depend on transcriptional activity of the host RNA polymerase II (Pol II) for their mobility, it was so far unclear whether Pol II is directly involved in repressing their activity.ResultsHere we show that plants defective in Pol II activity lose DNA methylation at repeat sequences and produce more extrachromosomal retrotransposon DNA upon stress in Arabidopsis and rice. We demonstrate that combined inhibition of both DNA methylation and Pol II activity leads to a strong stress-dependent mobilization of the heat responsive ONSEN retrotransposon in Arabidopsis seedlings. The progenies of these treated plants contain up to 75 new ONSEN insertions in their genome which are stably inherited over three generations of selfing. Repeated application of heat stress in progeny plants containing increased numbers of ONSEN copies does not result in increased activation of this transposon compared to control lines. Progenies with additional ONSEN copies show a broad panel of environment-dependent phenotypic diversity.ConclusionsWe demonstrate that Pol II acts at the root of transposon silencing. This is important because it suggests that Pol II can regulate the speed of plant evolution by fine-tuning the amplitude of transposon mobility. Our findings show that it is now possible to study induced transposon bursts in plants and unlock their use to induce epigenetic and genetic diversity for crop breeding.Electronic supplementary materialThe online version of this article (doi:10.1186/s13059-017-1265-4) contains supplementary material, which is available to authorized users.
“…eccDNA is a byproduct of the LTR retrotransposon life cycle [20]. Using mobilome sequencing, which comprises a specific amplification step of circular DNA followed by high-throughput sequencing to identify eccDNA derived from active LTR retrotransposons [2], we found that only ONSEN was activated by HS in combination with A&Z (Additional file 1: Figure S2). Confirming our qPCR data, more ONSEN -specific reads were detected in the presence of A and Z in the medium.Fig.…”
BackgroundRetrotransposons play a central role in plant evolution and could be a powerful endogenous source of genetic and epigenetic variability for crop breeding. To ensure genome integrity several silencing mechanisms have evolved to repress retrotransposon mobility. Even though retrotransposons fully depend on transcriptional activity of the host RNA polymerase II (Pol II) for their mobility, it was so far unclear whether Pol II is directly involved in repressing their activity.ResultsHere we show that plants defective in Pol II activity lose DNA methylation at repeat sequences and produce more extrachromosomal retrotransposon DNA upon stress in Arabidopsis and rice. We demonstrate that combined inhibition of both DNA methylation and Pol II activity leads to a strong stress-dependent mobilization of the heat responsive ONSEN retrotransposon in Arabidopsis seedlings. The progenies of these treated plants contain up to 75 new ONSEN insertions in their genome which are stably inherited over three generations of selfing. Repeated application of heat stress in progeny plants containing increased numbers of ONSEN copies does not result in increased activation of this transposon compared to control lines. Progenies with additional ONSEN copies show a broad panel of environment-dependent phenotypic diversity.ConclusionsWe demonstrate that Pol II acts at the root of transposon silencing. This is important because it suggests that Pol II can regulate the speed of plant evolution by fine-tuning the amplitude of transposon mobility. Our findings show that it is now possible to study induced transposon bursts in plants and unlock their use to induce epigenetic and genetic diversity for crop breeding.Electronic supplementary materialThe online version of this article (doi:10.1186/s13059-017-1265-4) contains supplementary material, which is available to authorized users.
“…cDNA can exist in both linear and circular forms, and circular forms were previously reported for EVADE and other members of the ATCOPIA93 family (Lanciano et al, 2017; Reinders et al, 2013). We observed outward-facing paired-end read alignments from Illumina VLP-seq samples mapping to ATCOPIA51 , ATCOPIA52 , EVADE , and ATGP3 , consistent with junction-crossing reads from circular templates (Fig.…”
15In Arabidopsis, LTR-retrotransposons are activated by mutations in the chromatin remodeler 16 DECREASE in DNA METHYLATION 1 (DDM1), giving rise to 21-22nt epigenetically 17 79 retrotransposons by stress, or by loss of histone methylation, also requires loss of 24nt small RNA 80 and RDR2/RNA polymerase IV (Ito et al., 2011; Mirouze et al., 2009). By contrast, in ddm1 81 mutants and wild-type pollen, most transposons are transcriptionally activated and 24nt siRNA 82 are partly replaced by 21-22nt easiRNA (Slotkin et al., 2009). In ddm1 mutants, easiRNA are 83 generated by RDR6 (Creasey et al., 2014; Nuthikattu et al., 2013) from the non-functional 84 ATHILA2 and ATHILA6 Ty3/gypsy retrotransposons but also from the functional, TY1/copia 85 element EVADE, and are triggered by diverse miRNA. In wild-type, retroelements generate 86 easiRNA only in pollen, where they are targeted at the PBS by miR845, and biogenesis occurs via 87 a non-canonical pathway (Borges et al., 2018).88 89 In order to develop a comprehensive catalog of functional retrotransposons in Arabidopsis, we 90 performed VLP DNA sequencing from ddm1 mutants, as well as genome-wide polysomal RNA 91 (translatome) and chromatin immunoprecipitation (ChIP) sequencing. VLP sequencing recovered 92 all known active retrotransposons in Arabidopsis, without the need for genome sequencing of 93 5 transposition events. Replication intermediates revealed profound differences between elements 94with multiple cPPT and high integration rates, and elements with no cPPT which preferentially 95 integrated into themselves ("suicidal" auto-integration within the VLP). We examined the roles of 96 easiRNA in retrotransposon control by investigating ddm1rdr6 double mutants (Creasey et al., 97 2014; Lippman et al., 2004; Vongs et al., 1993). We found that some retrotransposons are 98 regulated post-transcriptionally by RNA interference, while others are regulated at the 99 transcriptional level by histone H3 lysine-9 methylation guided by small RNA. We conclude that 100 easiRNA inhibits retrotransposition at multiple levels in the replication cycle and identify features 101 of active retrotransposons that promote activity and escape from silencing.
“…Commonly used procedure includes eccDNA enrichment and circular DNA analysis. The enrichment is involved in exonuclease digestion (such as Plasmid-Safe-ATP-dependent DNase and/or exonuclease V) to remove linear DNA and multiple displacement amplification (MDA) to preferentially amplify circular DNAs (1, 3, 5, 6, 16, 27, 36–38). This method has been proved to be effective for eccDNA detection and enrichment, especially when the initial DNA amount is very limited, for example the cell-free DNA (cfDNA) in the circulating system (27, 37).…”
Section: Methods For Identification and Enrichment Of Eccdnasmentioning
confidence: 99%
“…The eccDNAs are universally spread in eukaryotes, including plants (1, 2), yeasts (3, 4) and mammalians (5–7). Contrast to the more stable linear DNA on the chromosomes, the eccDNAs are extrachromosomally located, unstable, dynamic, and heterogeneously originated.…”
Extrachromosomal circular DNAs (eccDNAs) are circular DNAs that are originated from chromosomes, but are independent from chromosomal DNA. The eccDNAs are commonly found in various tissues and cell types, and in both normal and diseased conditions. Due to their highly heterogeneous origins and being widely spread in nearly all eukaryotes, the eccDNAs are believed to reflect the genome's plasticity and instability. With the assistance of next-generation sequencing, more eccDNAs have been characterized at the molecular level. Recently, eccDNAs have been reported as cell-free DNAs in the circulation system. Importantly, these circulating eccDNAs have shown some evidence with disease associations, suggesting their potential utility as a new type of biomarker for disease detection, treatment assessment and progress surveillance. However, many challenges need to be addressed before implementing the eccDNAs as a new source of genetic material for liquid biopsy.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
