Pathway conversion enables a double-lock mechanism to maintain DNA methylation and genome stability

The CMT2 and RNA-directed DNA methylation (RdDM) pathways have been proposed to separately maintain CHH methylation in specific regions of the Arabidopsis thaliana genome. Here, we show that dysfunction of the chromatin remodeler DDM1 causes hundreds of genomic regions to switch from CMT2 dependency to RdDM dependency in DNA methylation. These converted loci are enriched at the edge regions of long transposable elements (TEs). Furthermore, we found that dysfunction in both DDM1 and RdDM causes strong reactivation of TEs and a burst of TE transposition in the first generation of mutant plants, indicating that the DDM1 and RdDM pathways together are critical to maintaining TE repression and protecting genomic stability. Our findings reveal the existence of a pathway conversion–based backup mechanism to guarantee the maintenance of DNA methylation and genome integrity.

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The plant mobile domain proteins MAIN and MAIL1 interact with the phosphatase PP7L to regulate gene expression and silence transposable elements in Arabidopsis thaliana

10.1101/710905 ◽

2019 ◽

Author(s):

Melody Nicolau ◽

Nathalie Picault ◽

Julie Descombin ◽

Yasaman Jami-Alahmadi ◽

Suhua Feng ◽

...

Keyword(s):

Gene Expression ◽

Dna Methylation ◽

Arabidopsis Thaliana ◽

Transposable Elements ◽

Genome Stability ◽

Phylogenetic Analyses ◽

Dna Repeats ◽

Functional Protein ◽

Developmental Processes ◽

Phosphoprotein Phosphatase

ABSTRACTTransposable elements (TEs) are DNA repeats that must remain silenced to ensure cell integrity. Several epigenetic pathways including DNA methylation and histone modifications are involved in the silencing of TEs, and in the regulation of gene expression. In Arabidopsis thaliana, the TE-derived plant mobile domain (PMD) proteins have been involved in TE silencing, genome stability, and control of developmental processes. Using a forward genetic screen, we found that the PMD protein MAINTENANCE OF MERISTEMS (MAIN) acts synergistically and redundantly with DNA methylation to silence TEs. We found that MAIN and its close homolog MAIN-LIKE 1 (MAIL1) interact together, as well as with the phosphoprotein phosphatase (PPP) PP7-like (PP7L). Remarkably, main, mail1, pp7l single and mail1 pp7l double mutants display similar developmental phenotypes, and share common subsets of upregulated TEs and misregulated genes. Finally, phylogenetic analyses of PMD and PP7-type PPP domains among the Eudicot lineage suggest neo-association processes between the two protein domains to potentially generate new protein function. We propose that, through this interaction, the PMD and PPP domains may constitute a functional protein module required for the proper expression of a common set of genes, and for silencing of TEs.AUTHOR SUMMARYThe plant mobile domain (PMD) is a protein domain of unknown function that is widely spread in the angiosperm plants. Although most PMDs are associated with repeated DNA sequences called transposable elements (TEs), plants have domesticated the PMD to produce genic versions that play important roles within the cell. In Arabidopsis thaliana, MAINTENANCE OF MERISTEMS (MAIN) and MAIN-LIKE 1 (MAIL1) are genic PMDs that are involved in genome stability, developmental processes, and silencing of TEs. The mechanisms involving MAIN and MAIL1 in these cellular processes remain elusive. Here, we show that MAIN, MAIL1 and the phosphoprotein phosphatase (PPP) named PP7-like (PP7L) interact to form a protein complex that is required for the proper expression of genes, and the silencing of TEs. Phylogenetic analyses revealed that PMD and PP7-type PPP domains are evolutionary connected, and several plant species express proteins carrying both PMD and PPP domains. We propose that interaction of PMD and PPP domains would create a functional protein module involved in mechanisms regulating gene expression and repressing TEs.

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Distinct Responses of Arabidopsis Telomeres and Transposable Elements to Zebularine Exposure

International Journal of Molecular Sciences ◽

10.3390/ijms22010468 ◽

2021 ◽

Vol 22 (1) ◽

pp. 468

Author(s):

Klára Konečná ◽

Pavla Polanská Sováková ◽

Karin Anteková ◽

Jiří Fajkus ◽

Miloslava Fojtová

Keyword(s):

Transposable Elements ◽

Transcriptional Activation ◽

Genome Stability ◽

Cytosine Methylation ◽

Individual Variability ◽

Correlated Response ◽

Telomere Shortening ◽

Treatment Changes ◽

Telomere Homeostasis ◽

Genomic Regions

Involvement of epigenetic mechanisms in the regulation of telomeres and transposable elements (TEs), genomic regions with the protective and potentially detrimental function, respectively, has been frequently studied. Here, we analyzed telomere lengths in Arabidopsis thaliana plants of Columbia, Landsberg erecta and Wassilevskija ecotypes exposed repeatedly to the hypomethylation drug zebularine during germination. Shorter telomeres were detected in plants growing from seedlings germinated in the presence of zebularine with a progression in telomeric phenotype across generations, relatively high inter-individual variability, and diverse responses among ecotypes. Interestingly, the extent of telomere shortening in zebularine Columbia and Wassilevskija plants corresponded to the transcriptional activation of TEs, suggesting a correlated response of these genomic elements to the zebularine treatment. Changes in lengths of telomeres and levels of TE transcripts in leaves were not always correlated with a hypomethylation of cytosines located in these regions, indicating a cytosine methylation-independent level of their regulation. These observations, including differences among ecotypes together with distinct dynamics of the reversal of the disruption of telomere homeostasis and TEs transcriptional activation, reflect a complex involvement of epigenetic processes in the regulation of crucial genomic regions. Our results further demonstrate the ability of plant cells to cope with these changes without a critical loss of the genome stability.

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Hypomethylated poplars show higher tolerance to water deficit and highlight a dual role for DNA methylation in shoot meristem: regulation of stress response and genome integrity

10.1101/2020.04.16.045328 ◽

2020 ◽

Cited By ~ 1

Author(s):

M.D. Sow ◽

A-L. Le Gac ◽

R. Fichot ◽

S. Lanciano ◽

A. Delaunay ◽

...

Keyword(s):

Dna Methylation ◽

Stress Response ◽

Transposable Elements ◽

Water Deficit ◽

Shoot Apical Meristem ◽

Apical Meristem ◽

Shoot Meristem ◽

Genome Integrity ◽

Dependent Manner ◽

Altered Expression

AbstractAs fixed and long living organisms subjected to repeated environmental stresses, trees have developed mechanisms such as phenotypic plasticity that help them to cope with fluctuating environmental conditions. Here, we tested the role DNA methylation as a hub of integration, linking plasticity and physiological response to water deficit in the shoot apical meristem of the model tree poplar (Populus). Using a reverse genetic approach, we compared hypomethylated RNAi-ddm1 lines to wild-type trees for drought tolerance. An integrative analysis was realized with phytohormone balance, methylomes, transcriptomes and mobilomes.Hypomethylated lines were more tolerant when subjected to moderate water deficit and were intrinsically more tolerant to drought-induced cavitation. The alteration of the DDM1 machinery induced variation in DNA methylation in a cytosine context dependent manner, both in genes and transposable elements. Hypomethylated lines subjected to water deficit showed altered expression of genes involved in phytohormone pathways, such as salicylic acid and modified hormonal balance. Several transposable elements showed stress- and/or line-specific patterns of reactivation, and we could detect copy number variations for two of them in stressed ddm1 lines.Overall, our data highlight two major roles for DNA methylation in the shoot apical meristem: control of stress response and plasticity through transduction of hormone signaling and maintenance of genome integrity through the control of transposable elements.

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The RAD51 recombinase protects mitotic chromatin in human cells

Nature Communications ◽

10.1038/s41467-021-25643-y ◽

2021 ◽

Vol 12 (1) ◽

Cited By ~ 1

Author(s):

Isabel E. Wassing ◽

Emily Graham ◽

Xanita Saayman ◽

Lucia Rampazzo ◽

Christine Ralf ◽

...

Keyword(s):

Chromosome Segregation ◽

Genome Stability ◽

Spindle Assembly Checkpoint ◽

De Novo ◽

Genomic Stability ◽

Human Cells ◽

Genome Integrity ◽

Chromosomal Replication ◽

Anaphase Onset ◽

Mitotic Chromatin

AbstractThe RAD51 recombinase plays critical roles in safeguarding genome integrity, which is fundamentally important for all living cells. While interphase functions of RAD51 in maintaining genome stability are well-characterised, its role in mitosis remains contentious. In this study, we show that RAD51 protects under-replicated DNA in mitotic human cells and, in this way, promotes mitotic DNA synthesis (MiDAS) and successful chromosome segregation. In cells experiencing mild replication stress, MiDAS was detected irrespective of mitotically generated DNA damage. MiDAS broadly required de novo RAD51 recruitment to single-stranded DNA, which was supported by the phosphorylation of RAD51 by the key mitotic regulator Polo-like kinase 1. Importantly, acute inhibition of MiDAS delayed anaphase onset and induced centromere fragility, suggesting a mechanism that prevents the satisfaction of the spindle assembly checkpoint while chromosomal replication remains incomplete. This study hence identifies an unexpected function of RAD51 in promoting genomic stability in mitosis.

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Reciprocal interaction between SIRT6 and APC/C regulates genomic stability

Scientific Reports ◽

10.1038/s41598-021-93684-w ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Helin Wang ◽

Kangze Feng ◽

Qingtao Wang ◽

Haiteng Deng

Keyword(s):

Genome Stability ◽

Chromosome Instability ◽

Genomic Stability ◽

Centrosome Amplification ◽

Genome Integrity ◽

Anaphase Promoting Complex ◽

Mitotic Progression ◽

Master Regulator ◽

Reciprocal Interaction ◽

Proteasome Pathway

AbstractSIRT6 is an NAD+-dependent deacetylase that plays an important role in mitosis fidelity and genome stability. In the present study, we found that SIRT6 overexpression leads to mitosis defects and aneuploidy. We identified SIRT6 as a novel substrate of anaphase-promoting complex/cyclosome (APC/C), which is a master regulator of mitosis. Both CDH1 and CDC20, co-activators of APC/C, mediated SIRT6 degradation via the ubiquitination-proteasome pathway. Reciprocally, SIRT6 also deacetylated CDH1 at lysine K135 and promoted its degradation, resulting in an increase in APC/C-CDH1-targeted substrates, dysfunction in centrosome amplification, and chromosome instability. Our findings demonstrate the importance of SIRT6 for genome integrity during mitotic progression and reveal how SIRT6 and APC/C cooperate to drive mitosis.

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The Methylation Inhibitor 5-Aza-2′-Deoxycytidine Induces Genome-Wide Hypomethylation in Rice

10.21203/rs.3.rs-1147096/v1 ◽

2021 ◽

Author(s):

Shuo Liu ◽

Yu Bao ◽

Hui Deng ◽

Guanqing Liu ◽

Yangshuo Han ◽

...

Keyword(s):

Dna Methylation ◽

Genome Stability ◽

Epigenetic Modification ◽

Homozygous Mutation ◽

Global Methylation ◽

Genome Wide ◽

Methylation Inhibitor ◽

Dna Methylation Inhibitor ◽

Genomic Regions ◽

Genome Scale

Abstract DNA methylation is a conserved epigenetic modification which is vital for regulating gene expression and maintaining genome stability in both mammals and plants. Homozygous mutation of rice methyltransferase 1 (met1) gene can cause host death in rice, making it difficult to obtain plant material needed for hypomethylation research. To circumvent this challenge, the methylation inhibitor, 5-Aza-2′-deoxycytidine (AzaD), is used as a cytosine nucleoside analogue to reduce genome wide hypomethylation and is widely used in hypomethylation research. However, how AzaD affects plant methylation profiles at the genome scale is largely unknown. Here, we treated rice seedlings with AzaD and compared the AzaD treatment with osmet1-2 mutants, illustrating that there are similar CG hypomethylation and distribution throughout the whole genome. Along with global methylation loss class I transposable elements (TEs) which are farther from genes compared with class II TEs, were more significantly activated, and the RNA-directed DNA Methylation (RdDM) pathway was activated in specific genomic regions to compensate for severe CG loss. Overall, our results suggest that AzaD is an effective DNA methylation inhibitor that can influence genome wide methylation and cause a series of epigenetic variations.

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Changes in DNA Methylation in Arabidopsis thaliana Plants Exposed Over Multiple Generations to Gamma Radiation

Frontiers in Plant Science ◽

10.3389/fpls.2021.611783 ◽

2021 ◽

Vol 12 ◽

Author(s):

Pol Laanen ◽

Eline Saenen ◽

Mohamed Mysara ◽

Jorden Van de Walle ◽

May Van Hees ◽

...

Keyword(s):

Dna Methylation ◽

Arabidopsis Thaliana ◽

Transposable Elements ◽

Gamma Radiation ◽

Stress Responses ◽

Whole Genome ◽

Multiple Generations ◽

Three Generations ◽

Two Generations

Previous studies have found indications that exposure to ionising radiation (IR) results in DNA methylation changes in plants. However, this phenomenon is yet to be studied across multiple generations. Furthermore, the exact role of these changes in the IR-induced plant response is still far from understood. Here, we study the effect of gamma radiation on DNA methylation and its effect across generations in young Arabidopsis plants. A multigenerational set-up was used in which three generations (Parent, generation 1, and generation 2) of 7-day old Arabidopsis thaliana plants were exposed to either of the different radiation treatments (30, 60, 110, or 430 mGy/h) or to natural background radiation (control condition) for 14 days. The parental generation consisted of previously non-exposed plants, whereas generation 1 and generation 2 plants had already received a similar irradiation in the previous one or two generations, respectively. Directly after exposure the entire methylomes were analysed with UPLC-MS/MS to measure whole genome methylation levels. Whole genome bisulfite sequencing was used to identify differentially methylated regions (DMRs), including their methylation context in the three generations and this for three different radiation conditions (control, 30 mGy/h, and 110 mGy/h). Both intra- and intergenerational comparisons of the genes and transposable elements associated with the DMRs were made. Taking the methylation context into account, the highest number of changes were found for cytosines followed directly by guanine (CG methylation), whereas only limited changes in CHG methylation occurred and no changes in CHH methylation were observed. A clear increase in IR-induced DMRs was seen over the three generations that were exposed to the lowest dose rate, where generation 2 had a markedly higher number of DMRs than the previous two generations (Parent and generation 1). Counterintuitively, we did not see significant differences in the plants exposed to the highest dose rate. A large number of DMRs associated with transposable elements were found, the majority of them being hypermethylated, likely leading to more genetic stability. Next to that, a significant number of DMRs were associated with genes (either in their promoter-associated region or gene body). A functional analysis of these genes showed an enrichment for genes related to development as well as various stress responses, including DNA repair, RNA splicing, and (a)biotic stress responses. These observations indicate a role of DNA methylation in the regulation of these genes in response to IR exposure and shows a possible role for epigenetics in plant adaptation to IR over multiple generations.

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SMARCAD1 Mediated Active Replication Fork Stability Maintains Genome Integrity

10.1101/2020.10.05.326223 ◽

2020 ◽

Author(s):

Calvin Shun Yu Lo ◽

Marvin van Toorn ◽

Vincent Gaggioli ◽

Mariana Paes Dias ◽

Yifan Zhu ◽

...

Keyword(s):

Replication Fork ◽

Genome Stability ◽

Cellular Proliferation ◽

Genome Instability ◽

Genome Integrity ◽

Replication Forks ◽

Chromatin Remodeler ◽

Active Replication ◽

Fork Stalling ◽

Stalled Fork

ABSTRACTStalled fork protection pathway mediated by BRCA1/2 proteins is critical for replication fork stability that has implications in tumorigenesis. However, it is unclear if additional mechanisms are required to maintain replication fork stability. We describe a novel mechanism by which the chromatin remodeler SMARCAD1 stabilizes active replication forks that is essential for resistance towards replication poisons. We find that loss of SMARCAD1 results in toxic enrichment of 53BP1 at replication forks which mediates untimely dissociation of PCNA via the PCNA-unloader, ATAD5. Faster dissociation of PCNA causes frequent fork stalling, inefficient fork restart and accumulation of single-stranded DNA resulting in genome instability. Although, loss of 53BP1 in SMARCAD1 mutants restore PCNA levels, fork restart efficiency, genome stability and tolerance to replication poisons; this requires BRCA1 mediated fork protection. Interestingly, fork protection challenged BRCA1-deficient naïve- or PARPi-resistant tumors require SMARCAD1 mediated active fork stabilization to maintain unperturbed fork progression and cellular proliferation.

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Targeted epigenomic changes to the maize methylome resulting from tissue culture

10.1101/242081 ◽

2018 ◽

Cited By ~ 1

Author(s):

Zhaoxue Han ◽

Peter A. Crisp ◽

Scott Stelpflug ◽

Shawn M. Kaeppler ◽

Qing Li ◽

...

Keyword(s):

Dna Methylation ◽

Tissue Culture ◽

Regulation Of Gene Expression ◽

High Rate ◽

Genome Integrity ◽

Epigenetic State ◽

Context Specific ◽

Genomic Regions ◽

Methylation Patterns ◽

Gains And Losses

AbstractDNA methylation can contribute to the maintenance of genome integrity and regulation of gene expression. In most situations, DNA methylation patterns are inherited quite stably. However, changes in DNA methylation can occur at some loci as a result of tissue culture resulting in somaclonal variation. A sequence-capture bisulfite sequencing approach was implemented to monitor context-specific DNA methylation patterns in ~15Mb of the maize genome for a population of plants that had been regenerated from tissue culture. Plants that have been regenerated from tissue culture exhibit gains and losses of DNA methylation at a subset of genomic regions. There was evidence for a high rate of homozygous changes to DNA methylation levels that occur consistently in multiple independent tissue culture lines suggesting the existence of a targeted process for altering epigenetic state during tissue culture. The consistent changes induced by tissue culture include both gains and losses of DNA methylation and can affect CG, CHG or both contexts within a region. The majority of changes in DNA methylation exhibit stable inheritance although there is some evidence for stochastic reacquisition of the initial epigenetic state in some individuals. This study provides insights into the susceptibility of some loci and potential mechanisms that could contribute to altered DNA methylation and epigenetic state that occur during tissue culture in plant species.

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In plants distal regulatory sequences overlap with unmethylated rather than low-methylated regions, in contrast to mammals

10.1101/2020.03.24.005678 ◽

2020 ◽

Author(s):

Rurika Oka ◽

Mattijs Bliek ◽

Huub C.J. Hoefsloot ◽

Maike Stam

Keyword(s):

Gene Expression ◽

Dna Methylation ◽

Arabidopsis Thaliana ◽

Transcriptional Repression ◽

Genome Stability ◽

Regulation Of Gene Expression ◽

Regulatory Sequences ◽

Regulatory Regions ◽

Plant Genomes ◽

Underlying Mechanisms

AbstractBackgroundDNA methylation is an important factor in the regulation of gene expression and genome stability. High DNA methylation levels are associated with transcriptional repression. In mammalian systems, unmethylated, low methylated and fully methylated regions (UMRs, LMRs, and FMRs, respectively) can be distinguished. UMRs are associated with proximal regulatory regions, while LMRs are associated with distal regulatory regions. Although DNA methylation is mainly limited to the CG context in mammals, while it occurs in CG, CHG and CHH contexts in plants, UMRs and LMRs were expected to occupy similar genomic sequences in both mammals and plants.ResultsThis study investigated major model and crop plants such as Arabidopsis thaliana, tomato (Solanum lycopersicum), rice (Oryza sativa) and maize (Zea mays), and shows that plant genomes can also be subdivided in UMRs, LMRs and FMRs, but that LMRs are mainly present in the CHG context rather than the CG context. Strikingly, the identified CHG LMRs were enriched in transposable elements rather than regulatory regions. Maize candidate regulatory regions overlapped with UMRs. LMRs were enriched for heterochromatic histone modifications and depleted for DNase accessibility and H3K9 acetylation. CHG LMRs form a distinct, abundant cluster of loci, indicating they have a different role than FMRs.ConclusionsBoth mammalian and plant genomes can be segmented in three distinct classes of loci, UMRs, LMRs and FMRs, indicating similar underlying mechanisms. Unlike in mammals, distal regulatory sequences in plants appear to overlap with UMRs instead of LMRs. Our data indicate that LMRs in plants have a different function than those in mammals.

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