1,089 results on '"Jacobsen, Steven E."'
Search Results
2. Assessing the Biosynthetic Inventory of the Biocontrol Fungus Trichoderma afroharzianum T22.
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Han, Wenyu, Wu, Zhongshou, Zhong, Zhenhui, Williams, Jason, Jacobsen, Steven E, Sun, Zuodong, and Tang, Yi
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Hypocreales ,Biological Products ,Gene Expression Profiling ,Multigene Family ,Biosynthetic Pathways ,OSMAC ,biocontrol and biofertilizer fungi ,biosynthesis ,natural product ,secondary metabolite ,transcriptomics ,Chemical Sciences ,Agricultural and Veterinary Sciences ,Engineering ,Food Science - Abstract
Natural products biosynthesized from biocontrol fungi in the rhizosphere can have both beneficial and deleterious effects on plants. Herein, we performed a comprehensive analysis of natural product biosynthetic gene clusters (BGCs) from the widely used biocontrol fungus Trichoderma afroharzianum T22 (ThT22). This fungus encodes at least 64 BGCs, yet only seven compounds and four BGCs were previously characterized or mined. We correlated 21 BGCs of ThT22 with known primary and secondary metabolites through homologous BGC comparison and characterized one unknown BGC involved in the biosynthesis of eujavanicol A using heterologous expression. In addition, we performed untargeted transcriptomics and metabolic analysis to demonstrate the activation of silent ThT22 BGCs via the "one strain many compound" (OSMAC) approach. Collectively, our analysis showcases the biosynthetic capacity of ThT22 and paves the way for fully exploring the roles of natural products of ThT22.
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- 2023
3. MBD2 couples DNA methylation to transposable element silencing during male gametogenesis
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Wang, Shuya, Wang, Ming, Ichino, Lucia, Boone, Brandon A., Zhong, Zhenhui, Papareddy, Ranjith K., Lin, Evan K., Yun, Jaewon, Feng, Suhua, and Jacobsen, Steven E.
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- 2024
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4. A gene silencing screen uncovers diverse tools for targeted gene repression in Arabidopsis.
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Wang, Ming, Zhong, Zhenhui, Gallego-Bartolomé, Javier, Li, Zheng, Feng, Suhua, Kuo, Hsuan Yu, Kan, Ryan L, Lam, Hoiyan, Richey, John Curtis, Tang, Linli, Zhou, Jessica, Liu, Mukun, Jami-Alahmadi, Yasaman, Wohlschlegel, James, and Jacobsen, Steven E
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Arabidopsis ,Histones ,Arabidopsis Proteins ,Gene Expression ,Gene Expression Regulation ,Plant ,Gene Silencing ,Biotechnology ,Human Genome ,Genetics ,Plant Biology ,Crop and Pasture Production - Abstract
DNA methylation has been utilized for target gene silencing in plants. However, it is not well understood whether other silencing pathways can be also used to manipulate gene expression. Here we performed a gain-of-function screen for proteins that could silence a target gene when fused to an artificial zinc finger. We uncovered many proteins that suppressed gene expression through DNA methylation, histone H3K27me3 deposition, H3K4me3 demethylation, histone deacetylation, inhibition of RNA polymerase II transcription elongation or Ser-5 dephosphorylation. These proteins also silenced many other genes with different efficacies, and a machine learning model could accurately predict the efficacy of each silencer on the basis of various chromatin features of the target loci. Furthermore, some proteins were also able to target gene silencing when used in a dCas9-SunTag system. These results provide a more comprehensive understanding of epigenetic regulatory pathways in plants and provide an armament of tools for targeted gene manipulation.
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- 2023
5. Genome editing in plants using the compact editor CasΦ
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Li, Zheng, Zhong, Zhenhui, Wu, Zhongshou, Pausch, Patrick, Al-Shayeb, Basem, Amerasekera, Jasmine, Doudna, Jennifer A, and Jacobsen, Steven E
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Genetics ,Human Genome ,Biotechnology ,Gene Editing ,Arabidopsis ,CRISPR-Cas Systems ,Plants ,Genome ,Plant ,Transcription Factors ,Homeodomain Proteins ,Arabidopsis Proteins ,CRISPR-Cas? ,plant genome editing ,DNA methylation ,Cas? variants ,off-target editing ,CRISPR-CasΦ ,CasΦ variants - Abstract
Clustered regularly interspaced short palindromic repeats and CRISPR-associated proteins (CRISPR-Cas) systems have been developed as important tools for plant genome engineering. Here, we demonstrate that the hypercompact CasΦ nuclease is able to generate stably inherited gene edits in Arabidopsis, and that CasΦ guide RNAs can be expressed with either the Pol-III U6 promoter or a Pol-II promoter together with ribozyme mediated RNA processing. Using the Arabidopsis fwa epiallele, we show that CasΦ displays higher editing efficiency when the target locus is not DNA methylated, suggesting that CasΦ is sensitive to chromatin environment. Importantly, two CasΦ protein variants, vCasΦ and nCasΦ, both showed much higher editing efficiency relative to the wild-type CasΦ enzyme. Consistently, vCasΦ and nCasΦ yielded offspring plants with inherited edits at much higher rates compared to WTCasΦ. Extensive genomic analysis of gene edited plants showed no off-target editing, suggesting that CasΦ is highly specific. The hypercompact size, T-rich minimal protospacer adjacent motif (PAM), and wide range of working temperatures make CasΦ an excellent supplement to existing plant genome editing systems.
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- 2023
6. Improving cassava bacterial blight resistance by editing the epigenome
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Veley, Kira M, Elliott, Kiona, Jensen, Greg, Zhong, Zhenhui, Feng, Suhua, Yoder, Marisa, Gilbert, Kerrigan B, Berry, Jeffrey C, Lin, Zuh-Jyh Daniel, Ghoshal, Basudev, Gallego-Bartolomé, Javier, Norton, Joanna, Motomura-Wages, Sharon, Carrington, James C, Jacobsen, Steven E, and Bart, Rebecca S
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Human Genome ,Genetics ,Aetiology ,2.1 Biological and endogenous factors ,Infection ,Manihot ,Epigenome ,Xanthomonas ,Disease Resistance ,Transcription Factors ,Plant Diseases - Abstract
Pathogens rely on expression of host susceptibility (S) genes to promote infection and disease. As DNA methylation is an epigenetic modification that affects gene expression, blocking access to S genes through targeted methylation could increase disease resistance. Xanthomonas phaseoli pv. manihotis, the causal agent of cassava bacterial blight (CBB), uses transcription activator-like20 (TAL20) to induce expression of the S gene MeSWEET10a. In this work, we direct methylation to the TAL20 effector binding element within the MeSWEET10a promoter using a synthetic zinc-finger DNA binding domain fused to a component of the RNA-directed DNA methylation pathway. We demonstrate that this methylation prevents TAL20 binding, blocks transcriptional activation of MeSWEET10a in vivo and that these plants display decreased CBB symptoms while maintaining normal growth and development. This work therefore presents an epigenome editing approach useful for crop improvement.
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- 2023
7. MORC proteins regulate transcription factor binding by mediating chromatin compaction in active chromatin regions
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Zhong, Zhenhui, Xue, Yan, Harris, C Jake, Wang, Ming, Li, Zheng, Ke, Yunqing, Liu, Mukun, Zhou, Jessica, Jami-Alahmadi, Yasaman, Feng, Suhua, Wohlschlegel, James A, and Jacobsen, Steven E
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Biochemistry and Cell Biology ,Bioinformatics and Computational Biology ,Biological Sciences ,Genetics ,Plant Biology ,Underpinning research ,1.1 Normal biological development and functioning ,Generic health relevance ,Chromatin ,Arabidopsis ,Arabidopsis Proteins ,Gene Silencing ,Transcription Factors ,DNA Methylation ,Gene Expression Regulation ,Plant ,Microrchidia ,MORC ,Chromatin compaction ,TF binding ,Transcription regulation ,Environmental Sciences ,Information and Computing Sciences ,Bioinformatics - Abstract
BackgroundThe microrchidia (MORC) proteins are a family of evolutionarily conserved GHKL-type ATPases involved in chromatin compaction and gene silencing. Arabidopsis MORC proteins act in the RNA-directed DNA methylation (RdDM) pathway, where they act as molecular tethers to ensure the efficient establishment of RdDM and de novo gene silencing. However, MORC proteins also have RdDM-independent functions although their underlying mechanisms are unknown.ResultsIn this study, we examine MORC binding regions where RdDM does not occur in order to shed light on the RdDM-independent functions of MORC proteins. We find that MORC proteins compact chromatin and reduce DNA accessibility to transcription factors, thereby repressing gene expression. We also find that MORC-mediated repression of gene expression is particularly important under conditions of stress. MORC-regulated transcription factors can in some cases regulate their own transcription, resulting in feedback loops.ConclusionsOur findings provide insights into the molecular mechanisms of MORC-mediated chromatin compaction and transcription regulation.
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- 2023
8. Arabidopsis TRB proteins function in H3K4me3 demethylation by recruiting JMJ14
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Wang, Ming, Zhong, Zhenhui, Gallego-Bartolomé, Javier, Feng, Suhua, Shih, Yuan-Hsin, Liu, Mukun, Zhou, Jessica, Richey, John Curtis, Ng, Charmaine, Jami-Alahmadi, Yasaman, Wohlschlegel, James, Wu, Keqiang, and Jacobsen, Steven E
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Biological Sciences ,Genetics ,Biotechnology ,Histones ,Arabidopsis Proteins ,Arabidopsis ,Polycomb Repressive Complex 2 ,Telomere-Binding Proteins ,Demethylation ,Gene Expression Regulation ,Plant ,Polycomb-Group Proteins ,Jumonji Domain-Containing Histone Demethylases - Abstract
Arabidopsis telomeric repeat binding factors (TRBs) can bind telomeric DNA sequences to protect telomeres from degradation. TRBs can also recruit Polycomb Repressive Complex 2 (PRC2) to deposit tri-methylation of H3 lysine 27 (H3K27me3) over certain target loci. Here, we demonstrate that TRBs also associate and colocalize with JUMONJI14 (JMJ14) and trigger H3K4me3 demethylation at some loci. The trb1/2/3 triple mutant and the jmj14-1 mutant show an increased level of H3K4me3 over TRB and JMJ14 binding sites, resulting in up-regulation of their target genes. Furthermore, tethering TRBs to the promoter region of genes with an artificial zinc finger (TRB-ZF) successfully triggers target gene silencing, as well as H3K27me3 deposition, and H3K4me3 removal. Interestingly, JMJ14 is predominantly recruited to ZF off-target sites with low levels of H3K4me3, which is accompanied with TRB-ZFs triggered H3K4me3 removal at these loci. These results suggest that TRB proteins coordinate PRC2 and JMJ14 activities to repress target genes via H3K27me3 deposition and H3K4me3 removal.
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- 2023
9. A maternally programmed intergenerational mechanism enables male offspring to make piRNAs from Y-linked precursor RNAs in Drosophila
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Venkei, Zsolt G., Gainetdinov, Ildar, Bagci, Ayca, Starostik, Margaret R., Choi, Charlotte P., Fingerhut, Jaclyn M., Chen, Peiwei, Balsara, Chiraag, Whitfield, Troy W., Bell, George W., Feng, Suhua, Jacobsen, Steven E., Aravin, Alexei A., Kim, John K., Zamore, Phillip D., and Yamashita, Yukiko M.
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- 2023
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10. Single-nucleus RNA-seq reveals that MBD5, MBD6, and SILENZIO maintain silencing in the vegetative cell of developing pollen
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Ichino, Lucia, Picard, Colette L, Yun, Jaewon, Chotai, Meera, Wang, Shuya, Lin, Evan K, Papareddy, Ranjith K, Xue, Yan, and Jacobsen, Steven E
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Biochemistry and Cell Biology ,Bioinformatics and Computational Biology ,Genetics ,Biological Sciences ,Human Genome ,Underpinning research ,1.1 Normal biological development and functioning ,Generic health relevance ,RNA-Seq ,Arabidopsis ,Pollen ,DNA Transposable Elements ,Chromatin ,Transcription Factors ,DNA (Cytosine-5-)-Methyltransferases ,Arabidopsis Proteins ,DNA-Binding Proteins ,Highlights ,vegetative nuclei ,CP: Developmental biology ,CP: Plants ,DNA methylation ,MBD ,chromatin ,epigenetics ,male germline ,methyl readers ,pollen ,single-cell analysis ,transposable elements ,Medical Physiology ,Biological sciences - Abstract
Silencing of transposable elements (TEs) drives the evolution of numerous redundant mechanisms of transcriptional regulation. Arabidopsis MBD5, MBD6, and SILENZIO act as TE repressors downstream of DNA methylation. Here, we show, via single-nucleus RNA-seq of developing male gametophytes, that these repressors are critical for TE silencing in the pollen vegetative cell, a companion cell important for fertilization that undergoes chromatin decompaction. Instead, other silencing mutants (met1, ddm1, mom1, morc) show loss of silencing in all pollen nucleus types and somatic cells. We show that TEs repressed by MBD5/6 gain chromatin accessibility in wild-type vegetative nuclei despite remaining silent, suggesting that loss of DNA compaction makes them sensitive to loss of MBD5/6. Consistently, crossing mbd5/6 to histone 1 mutants, which have decondensed chromatin in leaves, reveals derepression of MBD5/6-dependent TEs in leaves. MBD5/6 and SILENZIO thus act as a silencing system particularly important when chromatin compaction is compromised.
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- 2022
11. H3.1K27me1 loss confers Arabidopsis resistance to Geminivirus by sequestering DNA repair proteins onto host genome
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Wang, Zhen, Castillo-González, Claudia M., Zhao, Changjiang, Tong, Chun-Yip, Li, Changhao, Zhong, Songxiao, Liu, Zhiyang, Xie, Kaili, Zhu, Jiaying, Wu, Zhongshou, Peng, Xu, Jacob, Yannick, Michaels, Scott D., Jacobsen, Steven E., and Zhang, Xiuren
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- 2023
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12. The MOM1 complex recruits the RdDM machinery via MORC6 to establish de novo DNA methylation
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Li, Zheng, Wang, Ming, Zhong, Zhenhui, Gallego-Bartolomé, Javier, Feng, Suhua, Jami-Alahmadi, Yasaman, Wang, Xinyi, Wohlschlegel, James, Bischof, Sylvain, Long, Jeff A., and Jacobsen, Steven E.
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- 2023
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13. CRISPR-CAS mediated transcriptional control and epi-mutagenesis
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Gardiner, Jason, Ghoshal, Basudev, Wang, Ming, and Jacobsen, Steven E
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Genetics ,Generic health relevance ,CRISPR-Cas Systems ,Mutagenesis ,Biological Sciences ,Agricultural and Veterinary Sciences ,Plant Biology & Botany - Abstract
Tools for sequence-specific DNA binding have opened the door to new approaches in investigating fundamental questions in biology and crop development. While there are several platforms to choose from, many of the recent advances in sequence-specific targeting tools are focused on developing Clustered Regularly Interspaced Short Palindromic Repeats- CRISPR Associated (CRISPR-Cas)-based systems. Using a catalytically inactive Cas protein (dCas), this system can act as a vector for different modular catalytic domains (effector domains) to control a gene's expression or alter epigenetic marks such as DNA methylation. Recent trends in developing CRISPR-dCas systems include creating versions that can target multiple copies of effector domains to a single site, targeting epigenetic changes that, in some cases, can be inherited to the next generation in the absence of the targeting construct, and combining effector domains and targeting strategies to create synergies that increase the functionality or efficiency of the system. This review summarizes and compares DNA targeting technologies, the effector domains used to target transcriptional control and epi-mutagenesis, and the different CRISPR-dCas systems used in plants.
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- 2022
14. The role of ATXR6 expression in modulating genome stability and transposable element repression in Arabidopsis
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Potok, Magdalena E, Zhong, Zhenhui, Picard, Colette L, Liu, Qikun, Do, Truman, Jacobsen, Cassidy E, Sakr, Ocean, Naranbaatar, Bilguudei, Thilakaratne, Ruwan, Khnkoyan, Zhanna, Purl, Megan, Cheng, Harrison, Vervaet, Helena, Feng, Suhua, Rayatpisheh, Shima, Wohlschlegel, James A, O’Malley, Ronan C, Ecker, Joseph R, and Jacobsen, Steven E
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Genetics ,Human Genome ,Biotechnology ,2.1 Biological and endogenous factors ,Aetiology ,Alleles ,Arabidopsis ,Arabidopsis Proteins ,DNA Transposable Elements ,Epigenesis ,Genetic ,Gene Expression Regulation ,Plant ,Genomic Instability ,Heterochromatin ,Histones ,Methyltransferases ,Mutation ,Phenotype ,Transcriptome ,H3K27me1 ,ATXR5/6 ,plant ,histone methyltransferase - Abstract
ARABIDOPSIS TRITHORAX-RELATED PROTEIN 5 (ATXR5) AND ATXR6 are required for the deposition of H3K27me1 and for maintaining genomic stability in Arabidopsis Reduction of ATXR5/6 activity results in activation of DNA damage response genes, along with tissue-specific derepression of transposable elements (TEs), chromocenter decompaction, and genomic instability characterized by accumulation of excess DNA from heterochromatin. How loss of ATXR5/6 and H3K27me1 leads to these phenotypes remains unclear. Here we provide extensive characterization of the atxr5/6 hypomorphic mutant by comprehensively examining gene expression and epigenetic changes in the mutant. We found that the tissue-specific phenotypes of TE derepression and excessive DNA in this atxr5/6 mutant correlated with residual ATXR6 expression from the hypomorphic ATXR6 allele. However, up-regulation of DNA damage genes occurred regardless of ATXR6 levels and thus appears to be a separable process. We also isolated an atxr6-null allele which showed that ATXR5 and ATXR6 are required for female germline development. Finally, we characterize three previously reported suppressors of the hypomorphic atxr5/6 mutant and show that these rescue atxr5/6 via distinct mechanisms, two of which involve increasing H3K27me1 levels.
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- 2022
15. Histone chaperone ASF1 mediates H3.3-H4 deposition in Arabidopsis
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Zhong, Zhenhui, Wang, Yafei, Wang, Ming, Yang, Fan, Thomas, Quentin Angelo, Xue, Yan, Zhang, Yaxin, Liu, Wanlu, Jami-Alahmadi, Yasaman, Xu, Linhao, Feng, Suhua, Marquardt, Sebastian, Wohlschlegel, James A, Ausin, Israel, and Jacobsen, Steven E
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Biochemistry and Cell Biology ,Genetics ,Biological Sciences ,Human Genome ,Biotechnology ,1.1 Normal biological development and functioning ,Underpinning research ,Generic health relevance ,Arabidopsis ,Cell Cycle Proteins ,Chromatin ,Chromatin Assembly Factor-1 ,Epigenesis ,Genetic ,Histone Chaperones ,Histones ,Molecular Chaperones ,Proteomics ,Arabidopsis Proteins - Abstract
Histone chaperones and chromatin remodelers control nucleosome dynamics, which are essential for transcription, replication, and DNA repair. The histone chaperone Anti-Silencing Factor 1 (ASF1) plays a central role in facilitating CAF-1-mediated replication-dependent H3.1 deposition and HIRA-mediated replication-independent H3.3 deposition in yeast and metazoans. Whether ASF1 function is evolutionarily conserved in plants is unknown. Here, we show that Arabidopsis ASF1 proteins display a preference for the HIRA complex. Simultaneous mutation of both Arabidopsis ASF1 genes caused a decrease in chromatin density and ectopic H3.1 occupancy at loci typically enriched with H3.3. Genetic, transcriptomic, and proteomic data indicate that ASF1 proteins strongly prefers the HIRA complex over CAF-1. asf1 mutants also displayed an increase in spurious Pol II transcriptional initiation and showed defects in the maintenance of gene body CG DNA methylation and in the distribution of histone modifications. Furthermore, ectopic targeting of ASF1 caused excessive histone deposition, less accessible chromatin, and gene silencing. These findings reveal the importance of ASF1-mediated histone deposition for proper epigenetic regulation of the genome.
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- 2022
16. The role of MORC3 in silencing transposable elements in mouse embryonic stem cells.
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Desai, Varsha P, Chouaref, Jihed, Wu, Haoyu, Pastor, William A, Kan, Ryan L, Oey, Harald M, Li, Zheng, Ho, Jamie, Vonk, Kelly KD, San Leon Granado, David, Christopher, Michael A, Clark, Amander T, Jacobsen, Steven E, and Daxinger, Lucia
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Chromatin regulators ,Endogenous retroviruses ,IAPs ,MORC3 ,MommeD screen ,Human Genome ,Stem Cell Research ,Genetics ,Biotechnology ,Stem Cell Research - Embryonic - Non-Human ,2.1 Biological and endogenous factors ,1.1 Normal biological development and functioning ,Cancer ,Generic health relevance - Abstract
BackgroundMicrorchidia proteins (MORCs) are involved in epigenetic gene silencing in a variety of eukaryotic organisms. Deletion of MORCs result in several developmental abnormalities and their dysregulation has been implicated in developmental disease and multiple cancers. Specifically, mammalian MORC3 mutations are associated with immune system defects and human cancers such as bladder, uterine, stomach, lung, and diffuse large B cell lymphomas. While previous studies have shown that MORC3 binds to H3K4me3 in vitro and overlaps with H3K4me3 ChIP-seq peaks in mouse embryonic stem cells, the mechanism by which MORC3 regulates gene expression is unknown.ResultsIn this study, we identified that mutation in Morc3 results in a suppressor of variegation phenotype in a Modifiers of murine metastable epialleles Dominant (MommeD) screen. We also find that MORC3 functions as an epigenetic silencer of transposable elements (TEs) in mouse embryonic stem cells (mESCs). Loss of Morc3 results in upregulation of TEs, specifically those belonging to the LTR class of retrotransposons also referred to as endogenous retroviruses (ERVs). Using ChIP-seq we found that MORC3, in addition to its known localization at H3K4me3 sites, also binds to ERVs, suggesting a direct role in regulating their expression. Previous studies have shown that these ERVs are marked by the repressive histone mark H3K9me3 which plays a key role in their silencing. However, we found that levels of H3K9me3 showed only minor losses in Morc3 mutant mES cells. Instead, we found that loss of Morc3 resulted in increased chromatin accessibility at ERVs as measured by ATAC-seq.ConclusionsOur results reveal MORC3 as a novel regulator of ERV silencing in mouse embryonic stem cells. The relatively minor changes of H3K9me3 in the Morc3 mutant suggests that MORC3 acts mainly downstream of, or in a parallel pathway with, the TRIM28/SETDB1 complex that deposits H3K9me3 at these loci. The increased chromatin accessibility of ERVs in the Morc3 mutant suggests that MORC3 may act at the level of chromatin compaction to effect TE silencing.
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- 2021
17. Arabidopsis MORC proteins function in the efficient establishment of RNA directed DNA methylation.
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Xue, Yan, Zhong, Zhenhui, Harris, C Jake, Gallego-Bartolomé, Javier, Wang, Ming, Picard, Colette, Cao, Xueshi, Hua, Shan, Kwok, Ivy, Feng, Suhua, Jami-Alahmadi, Yasaman, Sha, Jihui, Gardiner, Jason, Wohlschlegel, James, and Jacobsen, Steven E
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Arabidopsis ,Arabidopsis Proteins ,DNA Methylation ,Gene Expression Regulation ,Plant ,Gene Silencing ,Adenosine Triphosphatases - Abstract
The Microrchidia (MORC) family of ATPases are required for transposable element (TE) silencing and heterochromatin condensation in plants and animals, and C. elegans MORC-1 has been shown to topologically entrap and condense DNA. In Arabidopsis thaliana, mutation of MORCs has been shown to reactivate silent methylated genes and transposons and to decondense heterochromatic chromocenters, despite only minor changes in the maintenance of DNA methylation. Here we provide the first evidence localizing Arabidopsis MORC proteins to specific regions of chromatin and find that MORC4 and MORC7 are closely co-localized with sites of RNA-directed DNA methylation (RdDM). We further show that MORC7, when tethered to DNA by an artificial zinc finger, can facilitate the establishment of RdDM. Finally, we show that MORCs are required for the efficient RdDM mediated establishment of DNA methylation and silencing of a newly integrated FWA transgene, even though morc mutations have no effect on the maintenance of preexisting methylation at the endogenous FWA gene. We propose that MORCs function as a molecular tether in RdDM complexes to reinforce RdDM activity for methylation establishment. These findings have implications for MORC protein function in a variety of other eukaryotic organisms.
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- 2021
18. MBD5 and MBD6 couple DNA methylation to gene silencing through the J-domain protein SILENZIO
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Ichino, Lucia, Boone, Brandon A, Strauskulage, Luke, Harris, C Jake, Kaur, Gundeep, Gladstone, Matthew A, Tan, Maverick, Feng, Suhua, Jami-Alahmadi, Yasaman, Duttke, Sascha H, Wohlschlegel, James A, Cheng, Xiaodong, Redding, Sy, and Jacobsen, Steven E
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Biotechnology ,Human Genome ,Genetics ,1.1 Normal biological development and functioning ,Underpinning research ,2.1 Biological and endogenous factors ,Aetiology ,Generic health relevance ,General Science & Technology - Abstract
DNA methylation is associated with transcriptional repression of eukaryotic genes and transposons, but the downstream mechanism of gene silencing is largely unknown. Here we describe two Arabidopsis methyl-CpG binding domain proteins, MBD5 and MBD6, that are recruited to chromatin by recognition of CG methylation, and redundantly repress a subset of genes and transposons without affecting DNA methylation levels. These methyl-readers recruit a J-domain protein, SILENZIO, that acts as a transcriptional repressor in loss-of-function and gain-of-function experiments. J-domain proteins often serve as co-chaperones with HSP70s. Indeed, we found that SILENZIO's conserved J-domain motif was required for its interaction with HSP70s and for its silencing function. These results uncover an unprecedented role of a molecular chaperone J-domain protein in gene silencing downstream of DNA methylation.
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- 2021
19. CRISPR-based targeting of DNA methylation in Arabidopsis thaliana by a bacterial CG-specific DNA methyltransferase
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Ghoshal, Basudev, Picard, Colette L, Vong, Brandon, Feng, Suhua, and Jacobsen, Steven E
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Genetics ,Arabidopsis ,Bacterial Proteins ,CRISPR-Cas Systems ,DNA Methylation ,DNA ,Plant ,DNA-Cytosine Methylases ,Plants ,Genetically Modified ,Tenericutes ,DNA methylation ,CRISPR-Cas9 ,  ,SunTag - Abstract
CRISPR-based targeted modification of epigenetic marks such as DNA cytosine methylation is an important strategy to regulate the expression of genes and their associated phenotypes. Although plants have DNA methylation in all sequence contexts (CG, CHG, CHH, where H = A, T, C), methylation in the symmetric CG context is particularly important for gene silencing and is very efficiently maintained through mitotic and meiotic cell divisions. Tools that can directly add CG methylation to specific loci are therefore highly desirable but are currently lacking in plants. Here we have developed two CRISPR-based CG-specific targeted DNA methylation systems for plants using a variant of the bacterial CG-specific DNA methyltransferase MQ1 with reduced activity but high specificity. We demonstrate that the methylation added by MQ1 is highly target specific and can be heritably maintained in the absence of the effector. These tools should be valuable both in crop engineering and in plant genetic research.
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- 2021
20. Ectopic targeting of CG DNA methylation in Arabidopsis with the bacterial SssI methyltransferase.
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Liu, Wanlu, Gallego-Bartolomé, Javier, Zhou, Yuxing, Zhong, Zhenhui, Wang, Ming, Wongpalee, Somsakul Pop, Gardiner, Jason, Feng, Suhua, Kuo, Peggy Hsuanyu, and Jacobsen, Steven E
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Chromatin ,Spiroplasma ,Plants ,Genetically Modified ,Arabidopsis ,Bacterial Proteins ,Histones ,Arabidopsis Proteins ,DNA Methylation ,Gene Expression Regulation ,Plant ,DNA-Cytosine Methylases ,Chromatin Immunoprecipitation Sequencing ,RNA-Seq - Abstract
The ability to target epigenetic marks like DNA methylation to specific loci is important in both basic research and in crop plant engineering. However, heritability of targeted DNA methylation, how it impacts gene expression, and which epigenetic features are required for proper establishment are mostly unknown. Here, we show that targeting the CG-specific methyltransferase M.SssI with an artificial zinc finger protein can establish heritable CG methylation and silencing of a targeted locus in Arabidopsis. In addition, we observe highly heritable widespread ectopic CG methylation mainly over euchromatic regions. This hypermethylation shows little effect on transcription while it triggers a mild but significant reduction in the accumulation of H2A.Z and H3K27me3. Moreover, ectopic methylation occurs preferentially at less open chromatin that lacks positive histone marks. These results outline general principles of the heritability and interaction of CG methylation with other epigenomic features that should help guide future efforts to engineer epigenomes.
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- 2021
21. DNA methylation-linked chromatin accessibility affects genomic architecture in Arabidopsis
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Zhong, Zhenhui, Feng, Suhua, Duttke, Sascha H, Potok, Magdalena E, Zhang, Yiwei, Gallego-Bartolomé, Javier, Liu, Wanlu, and Jacobsen, Steven E
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Human Genome ,Genetics ,1.1 Normal biological development and functioning ,Underpinning research ,Generic health relevance ,Arabidopsis ,Chromatin ,DNA Methylation ,Genome ,Plant ,Mutation ,Transcription ,Genetic ,DNA methylation ,epigenetics ,chromatin accessibility - Abstract
DNA methylation is a major epigenetic modification found across species and has a profound impact on many biological processes. However, its influence on chromatin accessibility and higher-order genome organization remains unclear, particularly in plants. Here, we present genome-wide chromatin accessibility profiles of 18 Arabidopsis mutants that are deficient in CG, CHG, or CHH DNA methylation. We find that DNA methylation in all three sequence contexts impacts chromatin accessibility in heterochromatin. Many chromatin regions maintain inaccessibility when DNA methylation is lost in only one or two sequence contexts, and signatures of accessibility are particularly affected when DNA methylation is reduced in all contexts, suggesting an interplay between different types of DNA methylation. In addition, we found that increased chromatin accessibility was not always accompanied by increased transcription, suggesting that DNA methylation can directly impact chromatin structure by other mechanisms. We also observed that an increase in chromatin accessibility was accompanied by enhanced long-range chromatin interactions. Together, these results provide a valuable resource for chromatin architecture and DNA methylation analyses and uncover a pivotal role for methylation in the maintenance of heterochromatin inaccessibility.
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- 2021
22. Efficient and accurate determination of genome-wide DNA methylation patterns in Arabidopsis thaliana with enzymatic methyl sequencing
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Feng, Suhua, Zhong, Zhenhui, Wang, Ming, and Jacobsen, Steven E
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Biological Sciences ,Bioinformatics and Computational Biology ,Genetics ,Human Genome ,Arabidopsis ,DNA Methylation ,Epigenome ,Genome ,Plant ,Sensitivity and Specificity ,Sequence Analysis ,DNA ,DNA methylation ,Bisulfite sequencing ,WGBS ,EM-seq ,TET ,APOBEC ,Arabidopsis flowers ,Arabidopsis leaves - Abstract
Background5' methylation of cytosines in DNA molecules is an important epigenetic mark in eukaryotes. Bisulfite sequencing is the gold standard of DNA methylation detection, and whole-genome bisulfite sequencing (WGBS) has been widely used to detect methylation at single-nucleotide resolution on a genome-wide scale. However, sodium bisulfite is known to severely degrade DNA, which, in combination with biases introduced during PCR amplification, leads to unbalanced base representation in the final sequencing libraries. Enzymatic conversion of unmethylated cytosines to uracils can achieve the same end product for sequencing as does bisulfite treatment and does not affect the integrity of the DNA; enzymatic methylation sequencing may, thus, provide advantages over bisulfite sequencing.ResultsUsing an enzymatic methyl-seq (EM-seq) technique to selectively deaminate unmethylated cytosines to uracils, we generated and sequenced libraries based on different amounts of Arabidopsis input DNA and different numbers of PCR cycles, and compared these data to results from traditional whole-genome bisulfite sequencing. We found that EM-seq libraries were more consistent between replicates and had higher mapping and lower duplication rates, lower background noise, higher average coverage, and higher coverage of total cytosines. Differential methylation region (DMR) analysis showed that WGBS tended to over-estimate methylation levels especially in CHG and CHH contexts, whereas EM-seq detected higher CG methylation levels in certain highly methylated areas. These phenomena can be mostly explained by a correlation of WGBS methylation estimation with GC content and methylated cytosine density. We used EM-seq to compare methylation between leaves and flowers, and found that CHG methylation level is greatly elevated in flowers, especially in pericentromeric regions.ConclusionWe suggest that EM-seq is a more accurate and reliable approach than WGBS to detect methylation. Compared to WGBS, the results of EM-seq are less affected by differences in library preparation conditions or by the skewed base composition in the converted DNA. It may therefore be more desirable to use EM-seq in methylation studies.
- Published
- 2020
23. CRISPR-CasΦ from huge phages is a hypercompact genome editor
- Author
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Pausch, Patrick, Al-Shayeb, Basem, Bisom-Rapp, Ezra, Tsuchida, Connor A, Li, Zheng, Cress, Brady F, Knott, Gavin J, Jacobsen, Steven E, Banfield, Jillian F, and Doudna, Jennifer A
- Subjects
Biological Sciences ,Bioinformatics and Computational Biology ,Genetics ,Infectious Diseases ,Emerging Infectious Diseases ,Underpinning research ,1.1 Normal biological development and functioning ,Bacteriophages ,CRISPR-Cas Systems ,Clustered Regularly Interspaced Short Palindromic Repeats ,Gene Editing ,General Science & Technology - Abstract
CRISPR-Cas systems are found widely in prokaryotes, where they provide adaptive immunity against virus infection and plasmid transformation. We describe a minimal functional CRISPR-Cas system, comprising a single ~70-kilodalton protein, CasΦ, and a CRISPR array, encoded exclusively in the genomes of huge bacteriophages. CasΦ uses a single active site for both CRISPR RNA (crRNA) processing and crRNA-guided DNA cutting to target foreign nucleic acids. This hypercompact system is active in vitro and in human and plant cells with expanded target recognition capabilities relative to other CRISPR-Cas proteins. Useful for genome editing and DNA detection but with a molecular weight half that of Cas9 and Cas12a genome-editing enzymes, CasΦ offers advantages for cellular delivery that expand the genome editing toolbox.
- Published
- 2020
24. NAP1-RELATED PROTEIN1 and 2 negatively regulate H2A.Z abundance in chromatin in Arabidopsis.
- Author
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Wang, Yafei, Zhong, Zhenhui, Zhang, Yaxin, Xu, Linhao, Feng, Suhua, Rayatpisheh, Shima, Wohlschlegel, James A, Wang, Zonghua, Jacobsen, Steven E, and Ausin, Israel
- Abstract
In eukaryotes, DNA wraps around histones to form nucleosomes, which are compacted into chromatin. DNA-templated processes, including transcription, require chromatin disassembly and reassembly mediated by histone chaperones. Additionally, distinct histone variants can replace core histones to regulate chromatin structure and function. Although replacement of H2A with the evolutionarily conserved H2A.Z via the SWR1 histone chaperone complex has been extensively studied, in plants little is known about how a reduction of H2A.Z levels can be achieved. Here, we show that NRP proteins cause a decrease of H2A.Z-containing nucleosomes in Arabidopsis under standard growing conditions. nrp1-1 nrp2-2 double mutants show an over-accumulation of H2A.Z genome-wide, especially at heterochromatic regions normally H2A.Z-depleted in wild-type plants. Our work suggests that NRP proteins regulate gene expression by counteracting SWR1, thereby preventing excessive accumulation of H2A.Z.
- Published
- 2020
25. The characterization of Mediator 12 and 13 as conditional positive gene regulators in Arabidopsis.
- Author
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Liu, Qikun, Bischof, Sylvain, Harris, C Jake, Zhong, Zhenhui, Zhan, Lingyu, Nguyen, Calvin, Rashoff, Andrew, Barshop, William D, Sun, Fei, Feng, Suhua, Potok, Magdalena, Gallego-Bartolome, Javier, Zhai, Jixian, Wohlschlegel, James A, Carey, Michael F, Long, Jeffrey A, and Jacobsen, Steven E
- Abstract
Mediator 12 (MED12) and MED13 are components of the Mediator multi-protein complex, that facilitates the initial steps of gene transcription. Here, in an Arabidopsis mutant screen, we identify MED12 and MED13 as positive gene regulators, both of which contribute broadly to morc1 de-repressed gene expression. Both MED12 and MED13 are preferentially required for the expression of genes depleted in active chromatin marks, a chromatin signature shared with morc1 re-activated loci. We further discover that MED12 tends to interact with genes that are responsive to environmental stimuli, including light and radiation. We demonstrate that light-induced transient gene expression depends on MED12, and is accompanied by a concomitant increase in MED12 enrichment during induction. In contrast, the steady-state expression level of these genes show little dependence on MED12, suggesting that MED12 is primarily required to aid the expression of genes in transition from less-active to more active states.
- Published
- 2020
26. The plant mobile domain proteins MAIN and MAIL1 interact with the phosphatase PP7L to regulate gene expression and silence transposable elements in Arabidopsis thaliana.
- Author
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Nicolau, Melody, Picault, Nathalie, Descombin, Julie, Jami-Alahmadi, Yasaman, Feng, Suhua, Bucher, Etienne, Jacobsen, Steven E, Deragon, Jean-Marc, Wohlschlegel, James, and Moissiard, Guillaume
- Subjects
Heterochromatin ,Arabidopsis ,Nuclear Proteins ,Arabidopsis Proteins ,Repressor Proteins ,DNA Transposable Elements ,DNA Methylation ,Epigenesis ,Genetic ,Gene Expression Regulation ,Plant ,Gene Silencing ,Protein Binding ,Mutation ,Phosphoprotein Phosphatases ,Protein Domains ,Epigenesis ,Genetic ,Gene Expression Regulation ,Plant ,Genetics ,Developmental Biology - Abstract
Transposable 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.
- Published
- 2020
27. A kinesin Klp10A mediates cell cycle-dependent shuttling of Piwi between nucleus and nuage.
- Author
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Venkei, Zsolt G, Choi, Charlotte P, Feng, Suhua, Chen, Cuie, Jacobsen, Steven E, Kim, John K, and Yamashita, Yukiko M
- Subjects
Ovary ,Germ Cells ,Cell Nucleus ,Cytoplasm ,Animals ,Drosophila melanogaster ,Kinesin ,Cell Cycle Proteins ,Drosophila Proteins ,RNA ,Small Interfering ,DNA Transposable Elements ,Cell Cycle ,Cell Division ,Gene Silencing ,Female ,Male ,Argonaute Proteins ,RNA ,Small Interfering ,Genetics ,Developmental Biology - Abstract
The piRNA pathway protects germline genomes from selfish genetic elements (e.g. transposons) through their transcript cleavage in the cytoplasm and/or their transcriptional silencing in the nucleus. Here, we describe a mechanism by which the nuclear and cytoplasmic arms of the piRNA pathway are linked. We find that during mitosis of Drosophila spermatogonia, nuclear Piwi interacts with nuage, the compartment that mediates the cytoplasmic arm of the piRNA pathway. At the end of mitosis, Piwi leaves nuage to return to the nucleus. Dissociation of Piwi from nuage occurs at the depolymerizing microtubules of the central spindle, mediated by a microtubule-depolymerizing kinesin, Klp10A. Depletion of klp10A delays the return of Piwi to the nucleus and affects piRNA production, suggesting the role of nuclear-cytoplasmic communication in piRNA biogenesis. We propose that cell cycle-dependent communication between the nuclear and cytoplasmic arms of the piRNA pathway may play a previously unappreciated role in piRNA regulation.
- Published
- 2020
28. Promoter and Terminator Optimization for DNA Methylation Targeting in Arabidopsis
- Author
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Gardiner, Jason, Zhao, Jenny M, Chaffin, Kendall, and Jacobsen, Steven E
- Subjects
Biological Sciences ,Genetics ,Biotechnology ,Human Genome ,epigenetics ,DNA methylation ,targeting ,zinc finger ,Arabidopsis ,plant ,transcription ,promoter ,terminator ,RNA-directed DNA methylation ,silencing - Abstract
DNA methylation is an important epigenetic mark involved in gene regulation and silencing of transposable elements. The presence or absence of DNA methylation at specific sites can influence nearby gene expression and cause phenotypic changes that remain stable over generations. Recently, development of new technologies has enabled the targeted addition or removal of DNA methylation at specific sites of the genome. Of these new technologies, the targeting of the catalytic domain of Nicotiana tabacum DOMAINS REARRANGED METHYLTRANSFERASE 2 (ntDRM2cd) offers a promising tool for the addition of DNA methylation as it can directly methylate DNA. However, the methylation targeting efficiency of constructs using ntDRM2cd thus far has been relatively low. Previous studies have shown that the use of different promoters or terminators can greatly improve genome-editing efficiencies. In this study, we systematically survey a variety of promoter and terminator combinations to identify optimal combinations to use when targeting the addition of DNA methylation in Arabidopsis thaliana.
- Published
- 2020
29. Author response: H1 restricts euchromatin-associated methylation pathways from heterochromatic encroachment
- Author
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Harris, C. Jake, primary, Zhong, Zhenhui, additional, Ichino, Lucia, additional, Feng, Suhua, additional, and Jacobsen, Steven E., additional
- Published
- 2024
- Full Text
- View/download PDF
30. CryoEM structures of Arabidopsis DDR complexes involved in RNA-directed DNA methylation.
- Author
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Wongpalee, Somsakul Pop, Liu, Shiheng, Gallego-Bartolomé, Javier, Leitner, Alexander, Aebersold, Ruedi, Liu, Wanlu, Yen, Linda, Nohales, Maria A, Kuo, Peggy Hsuanyu, Vashisht, Ajay A, Wohlschlegel, James A, Feng, Suhua, Kay, Steve A, Zhou, Z Hong, and Jacobsen, Steven E
- Subjects
Arabidopsis ,Multiprotein Complexes ,DNA-Directed RNA Polymerases ,DNA-Binding Proteins ,Chromosomal Proteins ,Non-Histone ,Arabidopsis Proteins ,DNA ,Plant ,RNA ,Plant ,Cryoelectron Microscopy ,DNA Methylation ,Gene Expression Regulation ,Plant ,Protein Conformation ,Protein Binding ,Models ,Molecular ,Genetics ,1.1 Normal biological development and functioning ,Generic health relevance - Abstract
Transcription by RNA polymerase V (Pol V) in plants is required for RNA-directed DNA methylation, leading to transcriptional gene silencing. Global chromatin association of Pol V requires components of the DDR complex DRD1, DMS3 and RDM1, but the assembly process of this complex and the underlying mechanism for Pol V recruitment remain unknown. Here we show that all DDR complex components co-localize with Pol V, and we report the cryoEM structures of two complexes associated with Pol V recruitment-DR (DMS3-RDM1) and DDR' (DMS3-RDM1-DRD1 peptide), at 3.6 Å and 3.5 Å resolution, respectively. RDM1 dimerization at the center frames the assembly of the entire complex and mediates interactions between DMS3 and DRD1 with a stoichiometry of 1 DRD1:4 DMS3:2 RDM1. DRD1 binding to the DR complex induces a drastic movement of a DMS3 coiled-coil helix bundle. We hypothesize that both complexes are functional intermediates that mediate Pol V recruitment.
- Published
- 2019
31. Arabidopsis SWR1-associated protein methyl-CpG-binding domain 9 is required for histone H2A.Z deposition.
- Author
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Potok, Magdalena E, Wang, Yafei, Xu, Linhao, Zhong, Zhenhui, Liu, Wanlu, Feng, Suhua, Naranbaatar, Bilguudei, Rayatpisheh, Shima, Wang, Zonghua, Wohlschlegel, James A, Ausin, Israel, and Jacobsen, Steven E
- Subjects
Arabidopsis ,Microfilament Proteins ,Histones ,Arabidopsis Proteins ,Chromatin Assembly and Disassembly ,Gene Expression Regulation ,Plant ,Protein Binding ,Histone Acetyltransferases ,Gene Expression Regulation ,Plant - Abstract
Deposition of the histone variant H2A.Z by the SWI2/SNF2-Related 1 chromatin remodeling complex (SWR1-C) is important for gene regulation in eukaryotes, but the composition of the Arabidopsis SWR1-C has not been thoroughly characterized. Here, we aim to identify interacting partners of a conserved Arabidopsis SWR1 subunit ACTIN-RELATED PROTEIN 6 (ARP6). We isolate nine predicted components and identify additional interactors implicated in histone acetylation and chromatin biology. One of the interacting partners, methyl-CpG-binding domain 9 (MBD9), also strongly interacts with the Imitation SWItch (ISWI) chromatin remodeling complex. MBD9 is required for deposition of H2A.Z at a distinct subset of ARP6-dependent loci. MBD9 is preferentially bound to nucleosome-depleted regions at the 5' ends of genes containing high levels of activating histone marks. These data suggest that MBD9 is a SWR1-C interacting protein required for H2A.Z deposition at a subset of actively transcribing genes.
- Published
- 2019
32. Multi-level Modulation of Light Signaling by GIGANTEA Regulates Both the Output and Pace of the Circadian Clock.
- Author
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Nohales, Maria A, Liu, Wanlu, Duffy, Tomas, Nozue, Kazunari, Sawa, Mariko, Pruneda-Paz, Jose L, Maloof, Julin N, Jacobsen, Steven E, and Kay, Steve A
- Subjects
Arabidopsis ,Tobacco ,Plant Proteins ,Arabidopsis Proteins ,Signal Transduction ,Gene Expression Regulation ,Plant ,Circadian Rhythm ,Photoperiod ,Basic Helix-Loop-Helix Transcription Factors ,GIGANTEA ,circadian clock ,clock synchronization ,light input ,light signaling ,output ,transcriptional networks ,Sleep Research ,Underpinning research ,1.1 Normal biological development and functioning ,Biological Sciences ,Medical and Health Sciences ,Developmental Biology - Abstract
Integration of environmental signals with endogenous biological processes is essential for organisms to thrive in their natural environment. Being entrained by periodic environmental changes, the circadian clock incorporates external information to coordinate physiological processes, phasing them to the optimal time of the day and year. Here, we present a pivotal role for the clock component GIGANTEA (GI) as a genome-wide regulator of transcriptional networks mediating growth and adaptive processes in plants. We provide mechanistic details on how GI integrates endogenous timing with light signaling pathways through the global modulation of PHYTOCHROME-INTERACTING FACTORs (PIFs). Gating of the activity of these transcriptional regulators by GI directly affects a wide array of output rhythms, including photoperiodic growth. Furthermore, we uncover a role for PIFs in mediating light input to the circadian oscillator and show how their regulation by GI is required to set the pace of the clock in response to light-dark cycles.
- Published
- 2019
33. Genome mining and biosynthesis of a polyketide from a biofertilizer fungus that can facilitate reductive iron assimilation in plant
- Author
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Chen, Mengbin, Liu, Qikun, Gao, Shu-Shan, Young, Abbegayle E, Jacobsen, Steven E, and Tang, Yi
- Subjects
Plant Biology ,Biological Sciences ,Biotechnology ,Arabidopsis ,Iron ,Metabolic Networks and Pathways ,Methylation ,Molecular Chaperones ,Multigene Family ,Polyketide Synthases ,Polyketides ,Trichoderma ,iterative polyketide synthase ,biosynthesis ,natural products ,biofertilizer - Abstract
Fungi have the potential to produce a large repertoire of bioactive molecules, many of which can affect the growth and development of plants. Genomic survey of sequenced biofertilizer fungi showed many secondary metabolite gene clusters are anchored by iterative polyketide synthases (IPKSs), which are multidomain enzymes noted for generating diverse small molecules. Focusing on the biofertilizer Trichoderma harzianum t-22, we identified and characterized a cryptic IPKS-containing cluster that synthesizes tricholignan A, a redox-active ortho-hydroquinone. Tricholignan A is shown to reduce Fe(III) and may play a role in promoting plant growth under iron-deficient conditions. The construction of tricholignan by a pair of collaborating IPKSs was investigated using heterologous reconstitution and biochemical studies. A regioselective methylation step is shown to be a key step in formation of the ortho-hydroquinone. The responsible methyltransferase (MT) is fused with an N-terminal pseudo-acyl carrier protein (ψACP), in which the apo state of the ACP is essential for methylation of the growing polyketide chain. The ψACP is proposed to bind to the IPKS and enable the trans MT to access the growing polyketide. Our studies show that a genome-driven approach to discovering bioactive natural products from biofertilizer fungi can lead to unique compounds and biosynthetic knowledge.
- Published
- 2019
34. Co-targeting RNA Polymerases IV and V Promotes Efficient De Novo DNA Methylation in Arabidopsis.
- Author
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Gallego-Bartolomé, Javier, Liu, Wanlu, Kuo, Peggy Hsuanyu, Feng, Suhua, Ghoshal, Basudev, Gardiner, Jason, Zhao, Jenny Miao-Chi, Park, Soo Young, Chory, Joanne, and Jacobsen, Steven E
- Subjects
Arabidopsis ,Cytosine ,DNA-Directed RNA Polymerases ,RNA Polymerase II ,Arabidopsis Proteins ,RNA ,Small Interfering ,DNA ,RNA ,Plant ,DNA Methylation ,Gene Expression Regulation ,Plant ,Gene Expression Regulation ,Plant ,RNA ,Small Interfering ,Biological Sciences ,Medical and Health Sciences ,Developmental Biology - Abstract
The RNA-directed DNA methylation (RdDM) pathway in plants controls gene expression via cytosine DNA methylation. The ability to manipulate RdDM would shed light on the mechanisms and applications of DNA methylation to control gene expression. Here, we identified diverse RdDM proteins that are capable of targeting methylation and silencing in Arabidopsis when tethered to an artificial zinc finger (ZF-RdDM). We studied their order of action within the RdDM pathway by testing their ability to target methylation in different mutants. We also evaluated ectopic siRNA biogenesis, RNA polymerase V (Pol V) recruitment, targeted DNA methylation, and gene-expression changes at thousands of ZF-RdDM targets. We found that co-targeting both arms of the RdDM pathway, siRNA biogenesis and Pol V recruitment, dramatically enhanced targeted methylation. This work defines how RdDM components establish DNA methylation and enables new strategies for epigenetic gene regulation via targeted DNA methylation.
- Published
- 2019
35. A DNA methylation reader complex that enhances gene transcription
- Author
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Harris, C Jake, Scheibe, Marion, Wongpalee, Somsakul Pop, Liu, Wanlu, Cornett, Evan M, Vaughan, Robert M, Li, Xueqin, Chen, Wei, Xue, Yan, Zhong, Zhenhui, Yen, Linda, Barshop, William D, Rayatpisheh, Shima, Gallego-Bartolome, Javier, Groth, Martin, Wang, Zonghua, Wohlschlegel, James A, Du, Jiamu, Rothbart, Scott B, Butter, Falk, and Jacobsen, Steven E
- Subjects
Genetics ,1.1 Normal biological development and functioning ,Underpinning research ,Generic health relevance ,Arabidopsis ,Arabidopsis Proteins ,DNA Methylation ,Gene Expression Regulation ,Plant ,HSP40 Heat-Shock Proteins ,Histone-Lysine N-Methyltransferase ,Methyltransferases ,Protein Domains ,Transcription ,Genetic ,General Science & Technology - Abstract
DNA methylation generally functions as a repressive transcriptional signal, but it is also known to activate gene expression. In either case, the downstream factors remain largely unknown. By using comparative interactomics, we isolated proteins in Arabidopsis thaliana that associate with methylated DNA. Two SU(VAR)3-9 homologs, the transcriptional antisilencing factor SUVH1, and SUVH3, were among the methyl reader candidates. SUVH1 and SUVH3 bound methylated DNA in vitro, were associated with euchromatic methylation in vivo, and formed a complex with two DNAJ domain-containing homologs, DNAJ1 and DNAJ2. Ectopic recruitment of DNAJ1 enhanced gene transcription in plants, yeast, and mammals. Thus, the SUVH proteins bind to methylated DNA and recruit the DNAJ proteins to enhance proximal gene expression, thereby counteracting the repressive effects of transposon insertion near genes.
- Published
- 2018
36. The TFAP2C-Regulated OCT4 Naive Enhancer Is Involved in Human Germline Formation.
- Author
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Chen, Di, Liu, Wanlu, Zimmerman, Jill, Pastor, William A, Kim, Rachel, Hosohama, Linzi, Ho, Jamie, Aslanyan, Marianna, Gell, Joanna J, Jacobsen, Steven E, and Clark, Amander T
- Subjects
Germ Cells ,Chromatin ,Pluripotent Stem Cells ,Humans ,Gene Expression Regulation ,Developmental ,Female ,Male ,Transcription Factor AP-2 ,Octamer Transcription Factor-3 ,Kruppel-Like Transcription Factors ,Enhancer Elements ,Genetic ,Transcriptome ,Nucleotide Motifs ,OCT4 ,PGC ,PGCLC ,TFAP2C ,enhancer ,naive ,pluripotency ,Gene Expression Regulation ,Developmental ,Enhancer Elements ,Genetic ,Biochemistry and Cell Biology ,Medical Physiology - Abstract
Human primordial germ cells (hPGCs) are the first embryonic progenitors in the germ cell lineage, yet the molecular mechanisms required for hPGC formation are not well characterized. To identify regulatory regions in hPGC development, we used the assay for transposase-accessible chromatin using sequencing (ATAC-seq) to systematically characterize regions of open chromatin in hPGCs and hPGC-like cells (hPGCLCs) differentiated from human embryonic stem cells (hESCs). We discovered regions of open chromatin unique to hPGCs and hPGCLCs that significantly overlap with TFAP2C-bound enhancers identified in the naive ground state of pluripotency. Using CRISPR/Cas9, we show that deleting the TFAP2C-bound naive enhancer at the OCT4 locus (also called POU5F1) results in impaired OCT4 expression and a negative effect on hPGCLC identity.
- Published
- 2018
37. The impact and prospect of natural product discovery in agriculture
- Author
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Yan, Yan, Liu, Qikun, Jacobsen, Steven E, and Tang, Yi
- Subjects
Biochemistry and Cell Biology ,Biological Sciences ,Human Genome ,Biotechnology ,Genetics ,Complementary and Integrative Health ,Agriculture ,Biological Products ,Drug Resistance ,Microbial ,Fungicides ,Industrial ,Genome ,Microbial ,Genome ,Plant ,Metabolic Engineering ,Microorganisms ,Genetically-Modified ,Multigene Family ,Pesticides ,Plants ,Secondary Metabolism ,Developmental Biology ,Biochemistry and cell biology - Abstract
Natural products from plants and microorganisms are potent bioactive molecules that have been used in medicine, agriculture and cosmetics. Genomics and synthetic biology offer new tools to further explore their diversity for applications as fungicides or herbicides. [Image: see text]
- Published
- 2018
38. Mutations in DNA polymerase δ subunit 1 co-segregate with CMD2-type resistance to Cassava Mosaic Geminiviruses
- Author
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Lim, Yi-Wen, Mansfeld, Ben N., Schläpfer, Pascal, Gilbert, Kerrigan B., Narayanan, Narayanan N., Qi, Weihong, Wang, Qi, Zhong, Zhenhui, Boyher, Adam, Gehan, Jackson, Beyene, Getu, Lin, Zuh-Jyh Daniel, Esuma, Williams, Feng, Suhua, Chanez, Christelle, Eggenberger, Nadine, Adiga, Gerald, Alicai, Titus, Jacobsen, Steven E., Taylor, Nigel J., Gruissem, Wilhelm, and Bart, Rebecca S.
- Published
- 2022
- Full Text
- View/download PDF
39. Comprehensive identification of SWI/SNF complex subunits underpins deep eukaryotic ancestry and reveals new plant components
- Author
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Hernández-García, Jorge, Diego-Martin, Borja, Kuo, Peggy Hsuanyu, Jami-Alahmadi, Yasaman, Vashisht, Ajay A., Wohlschlegel, James, Jacobsen, Steven E., Blázquez, Miguel A., and Gallego-Bartolomé, Javier
- Published
- 2022
- Full Text
- View/download PDF
40. Mechanistic insights into plant SUVH family H3K9 methyltransferases and their binding to context-biased non-CG DNA methylation
- Author
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Li, Xueqin, Harris, C Jake, Zhong, Zhenhui, Chen, Wei, Liu, Rui, Jia, Bei, Wang, Zonghua, Li, Sisi, Jacobsen, Steven E, and Du, Jiamu
- Subjects
Genetics ,Human Genome ,Underpinning research ,1.1 Normal biological development and functioning ,Generic health relevance ,Arabidopsis ,Arabidopsis Proteins ,Base Sequence ,Crystallography ,X-Ray ,DNA ,DNA Methylation ,Histone-Lysine N-Methyltransferase ,Histones ,Lysine ,Methylation ,Models ,Molecular ,Nucleic Acid Conformation ,Plants ,Genetically Modified ,Protein Binding ,Protein Conformation ,DNA methylation ,H3K9me2 ,SUVH6 ,plant ,histone methyltransferase - Abstract
DNA methylation functions in gene silencing and the maintenance of genome integrity. In plants, non-CG DNA methylation is linked through a self-reinforcing loop with histone 3 lysine 9 dimethylation (H3K9me2). The plant-specific SUPPRESSOR OF VARIEGATION 3-9 HOMOLOG (SUVH) family H3K9 methyltransferases (MTases) bind to DNA methylation marks and catalyze H3K9 methylation. Here, we analyzed the structure and function of Arabidopsis thaliana SUVH6 to understand how this class of enzyme maintains methylation patterns in the genome. We reveal that SUVH6 has a distinct 5-methyl-dC (5mC) base-flipping mechanism involving a thumb loop element. Autoinhibition of H3 substrate entry is regulated by a SET domain loop, and a conformational transition in the post-SET domain upon cofactor binding may control catalysis. In vitro DNA binding and in vivo ChIP-seq data reveal that the different SUVH family H3K9 MTases have distinct DNA binding preferences, targeting H3K9 methylation to sites with different methylated DNA sequences, explaining the context biased non-CG DNA methylation in plants.
- Published
- 2018
41. Resistance-gene-directed discovery of a natural-product herbicide with a new mode of action.
- Author
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Yan, Yan, Liu, Qikun, Zang, Xin, Yuan, Shuguang, Bat-Erdene, Undramaa, Nguyen, Calvin, Gan, Jianhua, Zhou, Jiahai, Jacobsen, Steven E, and Tang, Yi
- Subjects
Plants ,Genetically Modified ,Arabidopsis ,Heterocyclic Compounds ,3-Ring ,Hydro-Lyases ,Plant Growth Regulators ,Biological Products ,Enzyme Inhibitors ,Herbicides ,Multigene Family ,Transgenes ,Models ,Molecular ,Herbicide Resistance ,Plants ,Genetically Modified ,Heterocyclic Compounds ,3-Ring ,Models ,Molecular ,General Science & Technology - Abstract
Bioactive natural products have evolved to inhibit specific cellular targets and have served as lead molecules for health and agricultural applications for the past century1-3. The post-genomics era has brought a renaissance in the discovery of natural products using synthetic-biology tools4-6. However, compared to traditional bioactivity-guided approaches, genome mining of natural products with specific and potent biological activities remains challenging4. Here we present the discovery and validation of a potent herbicide that targets a critical metabolic enzyme that is required for plant survival. Our approach is based on the co-clustering of a self-resistance gene in the natural-product biosynthesis gene cluster7-9, which provides insight into the potential biological activity of the encoded compound. We targeted dihydroxy-acid dehydratase in the branched-chain amino acid biosynthetic pathway in plants; the last step in this pathway is often targeted for herbicide development10. We show that the fungal sesquiterpenoid aspterric acid, which was discovered using the method described above, is a sub-micromolar inhibitor of dihydroxy-acid dehydratase that is effective as a herbicide in spray applications. The self-resistance gene astD was validated to be insensitive to aspterric acid and was deployed as a transgene in the establishment of plants that are resistant to aspterric acid. This herbicide-resistance gene combination complements the urgent ongoing efforts to overcome weed resistance11. Our discovery demonstrates the potential of using a resistance-gene-directed approach in the discovery of bioactive natural products.
- Published
- 2018
42. TFAP2C regulates transcription in human naive pluripotency by opening enhancers.
- Author
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Pastor, William A, Liu, Wanlu, Chen, Di, Ho, Jamie, Kim, Rachel, Hunt, Timothy J, Lukianchikov, Anastasia, Liu, Xiaodong, Polo, Jose M, Jacobsen, Steven E, and Clark, Amander T
- Subjects
Cells ,Cultured ,Chromatin ,Animals ,Humans ,Mice ,Signal Transduction ,Chromatin Assembly and Disassembly ,Species Specificity ,Transcription ,Genetic ,Gene Expression Regulation ,Developmental ,Binding Sites ,Protein Binding ,Cell Lineage ,Phenotype ,Transcription Factor AP-2 ,Octamer Transcription Factor-3 ,Enhancer Elements ,Genetic ,Nucleotide Motifs ,Human Embryonic Stem Cells ,Mouse Embryonic Stem Cells ,Cells ,Cultured ,Transcription ,Genetic ,Gene Expression Regulation ,Developmental ,Enhancer Elements ,Developmental Biology ,Biological Sciences ,Medical and Health Sciences - Abstract
Naive and primed pluripotent human embryonic stem cells bear transcriptional similarity to pre- and post-implantation epiblast and thus constitute a developmental model for understanding the pluripotent stages in human embryo development. To identify new transcription factors that differentially regulate the unique pluripotent stages, we mapped open chromatin using ATAC-seq and found enrichment of the activator protein-2 (AP2) transcription factor binding motif at naive-specific open chromatin. We determined that the AP2 family member TFAP2C is upregulated during primed to naive reversion and becomes widespread at naive-specific enhancers. TFAP2C functions to maintain pluripotency and repress neuroectodermal differentiation during the transition from primed to naive by facilitating the opening of enhancers proximal to pluripotency factors. Additionally, we identify a previously undiscovered naive-specific POU5F1 (OCT4) enhancer enriched for TFAP2C binding. Taken together, TFAP2C establishes and maintains naive human pluripotency and regulates OCT4 expression by mechanisms that are distinct from mouse.
- Published
- 2018
43. RNA-directed DNA methylation involves co-transcriptional small-RNA-guided slicing of polymerase V transcripts in Arabidopsis
- Author
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Liu, Wanlu, Duttke, Sascha H, Hetzel, Jonathan, Groth, Martin, Feng, Suhua, Gallego-Bartolome, Javier, Zhong, Zhenhui, Kuo, Hsuan Yu, Wang, Zonghua, Zhai, Jixian, Chory, Joanne, and Jacobsen, Steven E
- Subjects
Plant Biology ,Biological Sciences ,Bioinformatics and Computational Biology ,Genetics ,Biotechnology ,Human Genome ,Underpinning research ,1.1 Normal biological development and functioning ,Generic health relevance ,Arabidopsis ,Arabidopsis Proteins ,Argonaute Proteins ,DNA Methylation ,DNA-Directed RNA Polymerases ,Gene Silencing ,Genome-Wide Association Study ,RNA ,Guide ,Kinetoplastida ,RNA ,Small Nuclear ,Transcription ,Genetic ,Crop and Pasture Production ,Ecology ,Plant biology - Abstract
Small RNAs regulate chromatin modifications such as DNA methylation and gene silencing across eukaryotic genomes. In plants, RNA-directed DNA methylation (RdDM) requires 24-nucleotide small interfering RNAs (siRNAs) that bind to ARGONAUTE 4 (AGO4) and target genomic regions for silencing. RdDM also requires non-coding RNAs transcribed by RNA polymerase V (Pol V) that probably serve as scaffolds for binding of AGO4-siRNA complexes. Here, we used a modified global nuclear run-on protocol followed by deep sequencing to capture Pol V nascent transcripts genome-wide. We uncovered unique characteristics of Pol V RNAs, including a uracil (U) common at position 10. This uracil was complementary to the 5' adenine found in many AGO4-bound 24-nucleotide siRNAs and was eliminated in a siRNA-deficient mutant as well as in the ago4/6/9 triple mutant, suggesting that the +10 U signature is due to siRNA-mediated co-transcriptional slicing of Pol V transcripts. Expression of wild-type AGO4 in ago4/6/9 mutants was able to restore slicing of Pol V transcripts, but a catalytically inactive AGO4 mutant did not correct the slicing defect. We also found that Pol V transcript slicing required SUPPRESSOR OF TY INSERTION 5-LIKE (SPT5L), an elongation factor whose function is not well understood. These results highlight the importance of Pol V transcript slicing in RNA-mediated transcriptional gene silencing, which is a conserved process in many eukaryotes.
- Published
- 2018
44. Targeted DNA demethylation of the Arabidopsis genome using the human TET1 catalytic domain.
- Author
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Gallego-Bartolomé, Javier, Gardiner, Jason, Liu, Wanlu, Papikian, Ashot, Ghoshal, Basudev, Kuo, Hsuan Yu, Zhao, Jenny Miao-Chi, Segal, David J, and Jacobsen, Steven E
- Subjects
Humans ,Arabidopsis ,Flowers ,Mixed Function Oxygenases ,Homeodomain Proteins ,Proto-Oncogene Proteins ,Arabidopsis Proteins ,Transcription Factors ,DNA ,Plant ,DNA Transposable Elements ,DNA Methylation ,Epigenesis ,Genetic ,Gene Expression Regulation ,Plant ,Gene Silencing ,Catalytic Domain ,Zinc Fingers ,Mutation ,Genome ,Plant ,Promoter Regions ,Genetic ,CRISPR/dCas9 SunTag ,TET1 ,artificial zinc finger ,targeted demethylation ,DNA ,Plant ,Epigenesis ,Genetic ,Gene Expression Regulation ,Genome ,Promoter Regions - Abstract
DNA methylation is an important epigenetic modification involved in gene regulation and transposable element silencing. Changes in DNA methylation can be heritable and, thus, can lead to the formation of stable epialleles. A well-characterized example of a stable epiallele in plants is fwa, which consists of the loss of DNA cytosine methylation (5mC) in the promoter of the FLOWERING WAGENINGEN (FWA) gene, causing up-regulation of FWA and a heritable late-flowering phenotype. Here we demonstrate that a fusion between the catalytic domain of the human demethylase TEN-ELEVEN TRANSLOCATION1 (TET1cd) and an artificial zinc finger (ZF) designed to target the FWA promoter can cause highly efficient targeted demethylation, FWA up-regulation, and a heritable late-flowering phenotype. Additional ZF-TET1cd fusions designed to target methylated regions of the CACTA1 transposon also caused targeted demethylation and changes in expression. Finally, we have developed a CRISPR/dCas9-based targeted demethylation system using the TET1cd and a modified SunTag system. Similar to the ZF-TET1cd fusions, the SunTag-TET1cd system is able to target demethylation and activate gene expression when directed to the FWA or CACTA1 loci. Our study provides tools for targeted removal of 5mC at specific loci in the genome with high specificity and minimal off-target effects. These tools provide the opportunity to develop new epialleles for traits of interest, and to reactivate expression of previously silenced genes, transgenes, or transposons.
- Published
- 2018
45. Large-scale comparative epigenomics reveals hierarchical regulation of non-CG methylation in Arabidopsis
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Zhang, Yu, Harris, C Jake, Liu, Qikun, Liu, Wanlu, Ausin, Israel, Long, Yanping, Xiao, Lidan, Feng, Li, Chen, Xu, Xie, Yubin, Chen, Xinyuan, Zhan, Lingyu, Feng, Suhua, Li, Jingyi Jessica, Wang, Haifeng, Zhai, Jixian, and Jacobsen, Steven E
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Genetics ,Biotechnology ,Human Genome ,Generic health relevance ,Arabidopsis ,Cluster Analysis ,Computational Biology ,CpG Islands ,DNA Methylation ,Epigenesis ,Genetic ,Epigenomics ,Gene Expression Regulation ,Plant ,Gene Library ,Genome ,Plant ,Heterochromatin ,High-Throughput Nucleotide Sequencing ,Plants ,Genetically Modified ,Sequence Analysis ,DNA ,Sequence Analysis ,RNA ,Software ,epigenetics ,DNA methylation ,computational biology - Abstract
Genome-wide characterization by next-generation sequencing has greatly improved our understanding of the landscape of epigenetic modifications. Since 2008, whole-genome bisulfite sequencing (WGBS) has become the gold standard for DNA methylation analysis, and a tremendous amount of WGBS data has been generated by the research community. However, the systematic comparison of DNA methylation profiles to identify regulatory mechanisms has yet to be fully explored. Here we reprocessed the raw data of over 500 publicly available Arabidopsis WGBS libraries from various mutant backgrounds, tissue types, and stress treatments and also filtered them based on sequencing depth and efficiency of bisulfite conversion. This enabled us to identify high-confidence differentially methylated regions (hcDMRs) by comparing each test library to over 50 high-quality wild-type controls. We developed statistical and quantitative measurements to analyze the overlapping of DMRs and to cluster libraries based on their effect on DNA methylation. In addition to confirming existing relationships, we revealed unanticipated connections between well-known genes. For instance, MET1 and CMT3 were found to be required for the maintenance of asymmetric CHH methylation at nonoverlapping regions of CMT2 targeted heterochromatin. Our comparative methylome approach has established a framework for extracting biological insights via large-scale comparison of methylomes and can also be adopted for other genomics datasets.
- Published
- 2018
46. Germline competency of human embryonic stem cells depends on eomesodermin
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Chen, Di, Liu, Wanlu, Lukianchikov, Anastasia, Hancock, Grace V, Zimmerman, Jill, Lowe, Matthew G, Kim, Rachel, Galic, Zoran, Irie, Naoko, Surani, M Azim, Jacobsen, Steven E, and Clark, Amander T
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Biochemistry and Cell Biology ,Biomedical and Clinical Sciences ,Biological Sciences ,Stem Cell Research - Nonembryonic - Non-Human ,Stem Cell Research - Embryonic - Human ,Genetics ,Regenerative Medicine ,Stem Cell Research ,Biotechnology ,Underpinning research ,1.1 Normal biological development and functioning ,Generic health relevance ,CRISPR-Cas Systems ,Cell Differentiation ,Cell Line ,Female ,Gene Expression Regulation ,Developmental ,Germ Cells ,Human Embryonic Stem Cells ,Humans ,Male ,Sequence Analysis ,RNA ,Signal Transduction ,T-Box Domain Proteins ,human ,EOMES ,primordial germ cells ,embryonic stem cells ,Medical and Health Sciences ,Obstetrics & Reproductive Medicine ,Animal production ,Zoology ,Reproductive medicine - Abstract
In humans, germline competency and the specification of primordial germ cells (PGCs) are thought to occur in a restricted developmental window during early embryogenesis. Despite the importance of specifying the appropriate number of PGCs for human reproduction, the molecular mechanisms governing PGC formation remain largely unexplored. Here, we compared PGC-like cell (PGCLC) differentiation from 18 independently derived human embryonic stem cell (hESC) lines, and discovered that the expression of primitive streak genes were positively associated with hESC germline competency. Furthermore, we show that chemical inhibition of TGFβ and WNT signaling, which are required for primitive streak formation and CRISPR/Cas9 deletion of Eomesodermin (EOMES), significantly impacts PGCLC differentiation from hESCs. Taken together, our results suggest that human PGC formation involves signaling and transcriptional programs associated with somatic germ layer induction and expression of EOMES.
- Published
- 2017
47. The histone H3 variant H3.3 regulates gene body DNA methylation in Arabidopsis thaliana
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Wollmann, Heike, Stroud, Hume, Yelagandula, Ramesh, Tarutani, Yoshiaki, Jiang, Danhua, Jing, Li, Jamge, Bhagyshree, Takeuchi, Hidenori, Holec, Sarah, Nie, Xin, Kakutani, Tetsuji, Jacobsen, Steven E, and Berger, Frédéric
- Subjects
Plant Biology ,Biological Sciences ,Genetics ,Human Genome ,Prevention ,Biotechnology ,Underpinning research ,1.1 Normal biological development and functioning ,Generic health relevance ,Arabidopsis ,Arabidopsis Proteins ,Chromatin ,DNA (Cytosine-5-)-Methyltransferases ,DNA Methylation ,DNA ,Plant ,Epigenesis ,Genetic ,Genome ,Plant ,Histones ,Plant Proteins ,Transcription ,Genetic ,H3.3 ,Histone variants ,H2A.Z ,Linker histone H1 ,DNA methylation ,Environmental Sciences ,Information and Computing Sciences ,Bioinformatics - Abstract
BackgroundGene bodies of vertebrates and flowering plants are occupied by the histone variant H3.3 and DNA methylation. The origin and significance of these profiles remain largely unknown. DNA methylation and H3.3 enrichment profiles over gene bodies are correlated and both have a similar dependence on gene transcription levels. This suggests a mechanistic link between H3.3 and gene body methylation.ResultsWe engineered an H3.3 knockdown in Arabidopsis thaliana and observed transcription reduction that predominantly affects genes responsive to environmental cues. When H3.3 levels are reduced, gene bodies show a loss of DNA methylation correlated with transcription levels. To study the origin of changes in DNA methylation profiles when H3.3 levels are reduced, we examined genome-wide distributions of several histone H3 marks, H2A.Z, and linker histone H1. We report that in the absence of H3.3, H1 distribution increases in gene bodies in a transcription-dependent manner.ConclusionsWe propose that H3.3 prevents recruitment of H1, inhibiting H1's promotion of chromatin folding that restricts access to DNA methyltransferases responsible for gene body methylation. Thus, gene body methylation is likely shaped by H3.3 dynamics in conjunction with transcriptional activity.
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- 2017
48. MORC-1 Integrates Nuclear RNAi and Transgenerational Chromatin Architecture to Promote Germline Immortality
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Weiser, Natasha E, Yang, Danny X, Feng, Suhua, Kalinava, Natallia, Brown, Kristen C, Khanikar, Jayshree, Freeberg, Mallory A, Snyder, Martha J, Csankovszki, Györgyi, Chan, Raymond C, Gu, Sam G, Montgomery, Taiowa A, Jacobsen, Steven E, and Kim, John K
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Genetics ,Human Genome ,Contraception/Reproduction ,Biotechnology ,Underpinning research ,1.1 Normal biological development and functioning ,Generic health relevance ,Animals ,Caenorhabditis elegans ,Caenorhabditis elegans Proteins ,Cell Nucleus ,Chromatin ,Genome ,Germ Cells ,Heterochromatin ,Histones ,Inheritance Patterns ,Lysine ,Methylation ,Models ,Biological ,Mutation ,Nuclear Proteins ,RNA Interference ,RNA ,Small Interfering ,epigenetic inheritance ,germline ,heterochromatin ,histone modification ,microrchidia ,nuclear RNAi ,small RNA ,Biological Sciences ,Medical and Health Sciences ,Developmental Biology - Abstract
Germline-expressed endogenous small interfering RNAs (endo-siRNAs) transmit multigenerational epigenetic information to ensure fertility in subsequent generations. In Caenorhabditis elegans, nuclear RNAi ensures robust inheritance of endo-siRNAs and deposition of repressive H3K9me3 marks at target loci. How target silencing is maintained in subsequent generations is poorly understood. We discovered that morc-1 is essential for transgenerational fertility and acts as an effector of endo-siRNAs. Unexpectedly, morc-1 is dispensable for siRNA inheritance but is required for target silencing and maintenance of siRNA-dependent chromatin organization. A forward genetic screen identified mutations in met-1, which encodes an H3K36 methyltransferase, as potent suppressors of morc-1(-) and nuclear RNAi mutant phenotypes. Further analysis of nuclear RNAi and morc-1(-) mutants revealed a progressive, met-1-dependent enrichment of H3K36me3, suggesting that robust fertility requires repression of MET-1 activity at nuclear RNAi targets. Without MORC-1 and nuclear RNAi, MET-1-mediated encroachment of euchromatin leads to detrimental decondensation of germline chromatin and germline mortality.
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- 2017
49. Large-scale heterochromatin remodeling linked to overreplication-associated DNA damage
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Feng, Wei, Hale, Christopher J, Over, Ryan S, Cokus, Shawn J, Jacobsen, Steven E, and Michaels, Scott D
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Genetics ,Arabidopsis ,Arabidopsis Proteins ,Chromatin ,DNA Breaks ,Double-Stranded ,DNA Damage ,DNA Repair ,Gene Expression Regulation ,Plant ,Heterochromatin ,Histones ,Methyltransferases ,Mutation ,Phosphorylation ,ATXR5 ,ATXR6 ,H3K27me1 ,DNA repair ,DNA replication - Abstract
Previously, we have shown that loss of the histone 3 lysine 27 (H3K27) monomethyltransferases ARABIDOPSIS TRITHORAX-RELATED 5 (ATXR5) and ATXR6 (ATXR6) results in the overreplication of heterochromatin. Here we show that the overreplication results in DNA damage and extensive chromocenter remodeling into unique structures we have named "overreplication-associated centers" (RACs). RACs have a highly ordered structure with an outer layer of condensed heterochromatin, an inner layer enriched in the histone variant H2AX, and a low-density core containing foci of phosphorylated H2AX (a marker of double-strand breaks) and the DNA-repair enzyme RAD51. atxr5,6 mutants are strongly affected by mutations in DNA repair, such as ATM and ATR. Because of its dense packaging and repetitive DNA sequence, heterochromatin is a challenging environment in which to repair DNA damage. Previous work in animals has shown that heterochromatic breaks are translocated out of the heterochromatic domain for repair. Our results show that atxr5,6 mutants use a variation on this strategy for repairing heterochromatic DNA damage. Rather than being moved to adjacent euchromatic regions, as in animals, heterochromatin undergoes large-scale remodeling to create a compartment with low chromatin density.
- Published
- 2017
50. DNA methylome of the 20-gigabase Norway spruce genome
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Ausin, Israel, Feng, Suhua, Yu, Chaowei, Liu, Wanlu, Kuo, Hsuan Yu, Jacobsen, Elise L, Zhai, Jixian, Gallego-Bartolome, Javier, Wang, Lin, Egertsdotter, Ulrika, Street, Nathaniel R, Jacobsen, Steven E, and Wang, Haifeng
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Human Genome ,Biotechnology ,Genetics ,Underpinning research ,1.1 Normal biological development and functioning ,Base Sequence ,Cells ,Cultured ,Conserved Sequence ,Cycadopsida ,DNA Methylation ,DNA ,Plant ,Genome ,Plant ,Phylogeny ,Picea ,Plant Leaves ,gymnosperms ,Norway spruce ,DNA methylation ,small RNA ,genome size - Abstract
DNA methylation plays important roles in many biological processes, such as silencing of transposable elements, imprinting, and regulating gene expression. Many studies of DNA methylation have shown its essential roles in angiosperms (flowering plants). However, few studies have examined the roles and patterns of DNA methylation in gymnosperms. Here, we present genome-wide high coverage single-base resolution methylation maps of Norway spruce (Picea abies) from both needles and somatic embryogenesis culture cells via whole genome bisulfite sequencing. On average, DNA methylation levels of CG and CHG of Norway spruce were higher than most other plants studied. CHH methylation was found at a relatively low level; however, at least one copy of most of the RNA-directed DNA methylation pathway genes was found in Norway spruce, and CHH methylation was correlated with levels of siRNAs. In comparison with needles, somatic embryogenesis culture cells that are used for clonally propagating spruce trees showed lower levels of CG and CHG methylation but higher level of CHH methylation, suggesting that like in other species, these culture cells show abnormal methylation patterns.
- Published
- 2016
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