Dimethylation of histone H3 lysine 9 （H3K9me2） is an important epigenetic mark associated with transcription repression. Here, we identified PHF8, a JmjC-domain-containing protein, as a histone demethylase specific ...Dimethylation of histone H3 lysine 9 （H3K9me2） is an important epigenetic mark associated with transcription repression. Here, we identified PHF8, a JmjC-domain-containing protein, as a histone demethylase specific for this repressing mark. Recombinant full-length wild type protein could remove methylation from H3K9me2, but mutation of a conserved histidine to alanine H247A abolished the demethylase activity. Overexpressed exogenous PHF8 was colocalized with B23 staining. Endogenous PHF8 was also colocalized with B23 and fibrillarin, two well-established nucleolus proteins, suggesting that PHF8 is localized in the nucleolus and may regulate rRNA transcription. Indeed, PHF8 bound to the promoter region of the rDNA gene. Knockdown of PHF8 reduced the expression of rRNA, and overexpression of the gene resulted in upregulation of rRNA transcript. Concomitantly, H3K9me2 level was elevated in the promoter region of the rDNA gene in PHF8 knockdown cells and reduced significantly when the wild type but not the catalytically inactive H247A mutant PHF8 was overexpressed. Thus, our study identified a histone demethylase for H3K9me2 that regulates rRNA transcription.展开更多
RNA-directed DNA methylation(Rd DM) is a plant-specific de novo DNA methylation pathway,which has extensive cross-talk with histone modifications. Here, we report that the maize RdDM regulator SAWADEE HOMEODOMAIN HOMO...RNA-directed DNA methylation(Rd DM) is a plant-specific de novo DNA methylation pathway,which has extensive cross-talk with histone modifications. Here, we report that the maize RdDM regulator SAWADEE HOMEODOMAIN HOMOLOG 2(SHH2) is an H3 K9 me1 reader. Our structural studies reveal that H3 K9 me1 recognition is achieved by recognition of the methyl group via a classic aromatic cage and hydrogen-bonding and salt-bridge interactions with the free protons of the mono-methyllysine. The di-and tri-methylation states disrupt the polar interactions, decreasing the binding affinity. Our study reveals a monomethyllysine recognition mechanism which potentially links RdDM to H3 K9 me1 in maize.展开更多
DNA methylation,a conserved epigenetic mark,is critical for tuning temporal and spatial gene expression.The Arabidopsis thaliana DNA glycosylase/lyase REPRESSOR OF SILENCING 1(ROS1)initiates active DNA demethylation a...DNA methylation,a conserved epigenetic mark,is critical for tuning temporal and spatial gene expression.The Arabidopsis thaliana DNA glycosylase/lyase REPRESSOR OF SILENCING 1(ROS1)initiates active DNA demethylation and is required to prevent DNA hypermethylation at thousands of genomic loci.However,how ROS1 is recruited to specific loci is not well understood.Here,we report the discovery of Arabidopsis AGENET Domain Containing Protein 3(AGDP3)as a cellular factor that is required to prevent gene silencing and DNA hypermethylation.AGDP3 binds to H3K9me2 marks in its target DNA via its AGD12 cassette.Analysis of the crystal structure of the AGD12 cassette of AGDP3 in complex with an H3K9me2 peptide revealed that dimethylated H3 K9 and unmodified H3 K4 are specifically anchored into two different surface pockets.A histidine residue located in the methyllysine binding aromatic cage provides AGDP3 with pH-dependent H3K9me2 binding capacity.Our results uncover a molecular mechanism for the regulation of DNA demethylation by the gene silencing mark H3K9me2.展开更多
Serving as a host factor for human immunodeficiency virus(HIV)integration,LEDGF/p75 has been under extensive study as a potential target for therapy.However,as a highly conserved protein,its physiological function rem...Serving as a host factor for human immunodeficiency virus(HIV)integration,LEDGF/p75 has been under extensive study as a potential target for therapy.However,as a highly conserved protein,its physiological function remains to be thoroughly elucidated.Here,we characterize the molecular function of dP75,the Drosophila homolog of LEDGF/p75,during oogenesis.dP75 binds to transcriptionally active chromatin with its PWWP domain.The C-terminus integrase-binding domain-containing region of dP75 physically interacts with the histone kinase Jil-1 and stabilizes it in vivo.Together with Jil-1,dP75 prevents the spreading of the heterochromatin mark-H3 K9 me2-onto genes required for oogenesis and piRNA production.Without dP75,ectopical silencing of these genes disrupts oogenesis,activates transposons,and causes animal sterility.We propose that dP75,the homolog of an HIV host factor in Drosophila,partners with and stabilizes Jil-1 to ensure gene expression during oogenesis by preventing ectopic heterochromatin spreading.展开更多
The human genome can be demarcated into domains based on distinct epigenetic states.The trimethylation of histone H3 lysine 9(H3K9me3)is essential for the formation of constitutive heterochromatin nanodomains.However,...The human genome can be demarcated into domains based on distinct epigenetic states.The trimethylation of histone H3 lysine 9(H3K9me3)is essential for the formation of constitutive heterochromatin nanodomains.However,the extent to which genomic regions require specific densities or degrees of H3K9me3 for stable interactions remains unclear.Here,we utilize CRISPR-based DNA imaging to investigate the role of endogenous or ectopic H3K9me3 in chromatin dynamics and genomic interactions.We select three loci(IDR3,TCF3,and PR1)with distinct levels of H3K9me3 to examine the genomic interactions and association with endogenous Heterochromatin Protein 1(HP1α)condensates.Our results demonstrate a positive correlation between the levels of H3K9me3 at the loci and their association with HP1αcondensates.By dual-color labeling and long-term tracking of IDR3 and PR1 loci,we find a periodical association between the two ranging from one to three hours.Epigenetic perturbation-induced Genome organization(EpiGo)-KRAB introduces∼20 kilobases of H3K9me3 at the TCF3 locus,which is sufficient to establish a stable association between TCF3 and HP1αcondensates.In addition,EpiGo-mediated H3K9me3 also leads to stable genomic interaction between IDR3 and TCF3.Briefly,these data suggest that the density of H3K9me3 could dictate the stability of interactions between genomic loci and HP1αcondensates.展开更多
In the nucleus, chromatin is folded into hierarchical architecture that is tightly linked to various nuclear functions. However, the underlying molecular mechanisms that confer these architectures remain incompletely ...In the nucleus, chromatin is folded into hierarchical architecture that is tightly linked to various nuclear functions. However, the underlying molecular mechanisms that confer these architectures remain incompletely understood. Here, we investigated the functional roles of H3 lysine 9 dimethylation(H3 K9 me2), one of the abundant histone modifications, in three-dimensional(3 D)genome organization. Unlike in mouse embryonic stem cells, inhibition of methyltransferases G9 a and GLP in differentiated cells eliminated H3 K9 me2 predominantly at A-type(active) genomic compartments, and the level of residual H3 K9 me2 modifications was strongly associated with B-type(inactive) genomic compartments. Furthermore, chemical inhibition of G9 a/GLP in mouse hepatocytes led to decreased chromatin-nuclear lamina interactions mainly at G9 a/GLP-sensitive regions, increased degree of genomic compartmentalization, and up-regulation of hundreds of genes that were associated with alterations of the 3 D chromatin. Collectively, our data demonstrated essential roles of H3 K9 me2 in 3 D genome organization.展开更多
基金Acknowledgments We thank the cell biology core facility for confocal study. The PHF8 antibody was kindly provided by Dr Jiemin Wong （East China Normal University）. This work was supported by the National Basic Research Program of China （2007CB947900, 2010CB529705, 2007CB947100）, the Chinese Academy of Sci- ences （KSCX2-YW-R-04, KSCX2-YW-R-I 11）, the National Natural Science Foundation of China （30870538, 90919026）, Postdoctoral fellowship （20090460670）, and the Council of Shanghai Municipal Government for Science and Technology.
文摘Dimethylation of histone H3 lysine 9 （H3K9me2） is an important epigenetic mark associated with transcription repression. Here, we identified PHF8, a JmjC-domain-containing protein, as a histone demethylase specific for this repressing mark. Recombinant full-length wild type protein could remove methylation from H3K9me2, but mutation of a conserved histidine to alanine H247A abolished the demethylase activity. Overexpressed exogenous PHF8 was colocalized with B23 staining. Endogenous PHF8 was also colocalized with B23 and fibrillarin, two well-established nucleolus proteins, suggesting that PHF8 is localized in the nucleolus and may regulate rRNA transcription. Indeed, PHF8 bound to the promoter region of the rDNA gene. Knockdown of PHF8 reduced the expression of rRNA, and overexpression of the gene resulted in upregulation of rRNA transcript. Concomitantly, H3K9me2 level was elevated in the promoter region of the rDNA gene in PHF8 knockdown cells and reduced significantly when the wild type but not the catalytically inactive H247A mutant PHF8 was overexpressed. Thus, our study identified a histone demethylase for H3K9me2 that regulates rRNA transcription.
基金supported by National Natural Science Foundation of China(31770782)the Ministry of Science and Technology of China(2016YFA0503200)+1 种基金the Shenzhen Science and Technology Program(JCYJ20200109110403829 and KQTD20190929173906742)Key Laboratory of Molecular Design for Plant Cell Factory of Guangdong Higher Education Institutes(2019KSYS006)to J.D.
文摘RNA-directed DNA methylation(Rd DM) is a plant-specific de novo DNA methylation pathway,which has extensive cross-talk with histone modifications. Here, we report that the maize RdDM regulator SAWADEE HOMEODOMAIN HOMOLOG 2(SHH2) is an H3 K9 me1 reader. Our structural studies reveal that H3 K9 me1 recognition is achieved by recognition of the methyl group via a classic aromatic cage and hydrogen-bonding and salt-bridge interactions with the free protons of the mono-methyllysine. The di-and tri-methylation states disrupt the polar interactions, decreasing the binding affinity. Our study reveals a monomethyllysine recognition mechanism which potentially links RdDM to H3 K9 me1 in maize.
基金the Chinese Academy of Sciences and the National Natural Science Foundation of China(31970580)to M.L.the National Key R&D Program(2016YFA0503200)+1 种基金Shenzhen Science and Technology Program(JCYJ20200109110403829 and KQTD20190929173906742)Key Laboratory of Molecular Design for Plant Cell Factory of Guangdong Higher Education Institutes(2019KSYS006)to J.D.
文摘DNA methylation,a conserved epigenetic mark,is critical for tuning temporal and spatial gene expression.The Arabidopsis thaliana DNA glycosylase/lyase REPRESSOR OF SILENCING 1(ROS1)initiates active DNA demethylation and is required to prevent DNA hypermethylation at thousands of genomic loci.However,how ROS1 is recruited to specific loci is not well understood.Here,we report the discovery of Arabidopsis AGENET Domain Containing Protein 3(AGDP3)as a cellular factor that is required to prevent gene silencing and DNA hypermethylation.AGDP3 binds to H3K9me2 marks in its target DNA via its AGD12 cassette.Analysis of the crystal structure of the AGD12 cassette of AGDP3 in complex with an H3K9me2 peptide revealed that dimethylated H3 K9 and unmodified H3 K4 are specifically anchored into two different surface pockets.A histidine residue located in the methyllysine binding aromatic cage provides AGDP3 with pH-dependent H3K9me2 binding capacity.Our results uncover a molecular mechanism for the regulation of DNA demethylation by the gene silencing mark H3K9me2.
基金supported by the grant from the NIH to Z.Z.(DP5OD021355)the National Natural Science Foundation of China(91940302 and 31870741 to Y.H.)。
文摘Serving as a host factor for human immunodeficiency virus(HIV)integration,LEDGF/p75 has been under extensive study as a potential target for therapy.However,as a highly conserved protein,its physiological function remains to be thoroughly elucidated.Here,we characterize the molecular function of dP75,the Drosophila homolog of LEDGF/p75,during oogenesis.dP75 binds to transcriptionally active chromatin with its PWWP domain.The C-terminus integrase-binding domain-containing region of dP75 physically interacts with the histone kinase Jil-1 and stabilizes it in vivo.Together with Jil-1,dP75 prevents the spreading of the heterochromatin mark-H3 K9 me2-onto genes required for oogenesis and piRNA production.Without dP75,ectopical silencing of these genes disrupts oogenesis,activates transposons,and causes animal sterility.We propose that dP75,the homolog of an HIV host factor in Drosophila,partners with and stabilizes Jil-1 to ensure gene expression during oogenesis by preventing ectopic heterochromatin spreading.
基金This work was funded by the National Natural Science Foundation of China(No.31970591 to H.Ma),the Shanghai Pujiang Program(19PJ1408000 to H.Ma)the Shanghai Science and Technology Innovation Action Plan(21JC1404800 to H.Ma)We thank Luke Lavis(Janelia Research Campus,Howard Hughes Medical Institute,Ashburn,VA,USA)for providing the HaloTag JF-549.U2OS Genomic DNA was a gift from Xingxu Huang.We thank Pengwei Zhang and Shuangli Zhang for their help with cell sorting.
文摘The human genome can be demarcated into domains based on distinct epigenetic states.The trimethylation of histone H3 lysine 9(H3K9me3)is essential for the formation of constitutive heterochromatin nanodomains.However,the extent to which genomic regions require specific densities or degrees of H3K9me3 for stable interactions remains unclear.Here,we utilize CRISPR-based DNA imaging to investigate the role of endogenous or ectopic H3K9me3 in chromatin dynamics and genomic interactions.We select three loci(IDR3,TCF3,and PR1)with distinct levels of H3K9me3 to examine the genomic interactions and association with endogenous Heterochromatin Protein 1(HP1α)condensates.Our results demonstrate a positive correlation between the levels of H3K9me3 at the loci and their association with HP1αcondensates.By dual-color labeling and long-term tracking of IDR3 and PR1 loci,we find a periodical association between the two ranging from one to three hours.Epigenetic perturbation-induced Genome organization(EpiGo)-KRAB introduces∼20 kilobases of H3K9me3 at the TCF3 locus,which is sufficient to establish a stable association between TCF3 and HP1αcondensates.In addition,EpiGo-mediated H3K9me3 also leads to stable genomic interaction between IDR3 and TCF3.Briefly,these data suggest that the density of H3K9me3 could dictate the stability of interactions between genomic loci and HP1αcondensates.
基金supported by the National Key R&D Program of China(Grant No.2018YFC1003500 awarded to BW)the National Natural Science Foundation of China(Grant No.31771435 awarded to BW)
文摘In the nucleus, chromatin is folded into hierarchical architecture that is tightly linked to various nuclear functions. However, the underlying molecular mechanisms that confer these architectures remain incompletely understood. Here, we investigated the functional roles of H3 lysine 9 dimethylation(H3 K9 me2), one of the abundant histone modifications, in three-dimensional(3 D)genome organization. Unlike in mouse embryonic stem cells, inhibition of methyltransferases G9 a and GLP in differentiated cells eliminated H3 K9 me2 predominantly at A-type(active) genomic compartments, and the level of residual H3 K9 me2 modifications was strongly associated with B-type(inactive) genomic compartments. Furthermore, chemical inhibition of G9 a/GLP in mouse hepatocytes led to decreased chromatin-nuclear lamina interactions mainly at G9 a/GLP-sensitive regions, increased degree of genomic compartmentalization, and up-regulation of hundreds of genes that were associated with alterations of the 3 D chromatin. Collectively, our data demonstrated essential roles of H3 K9 me2 in 3 D genome organization.
