RNA介导的DNA甲基化作用(RNA-directed DNA Methylation,RdDM)是首次在植物中发现的基因组表观修饰现象,RdDM通过RNA-DNA序列相互作用直接导致DNA甲基化。植物中的RdDM和siRNA介导的mRNA降解现象,都是通过RNA使序列特异性基因发生沉默,...RNA介导的DNA甲基化作用(RNA-directed DNA Methylation,RdDM)是首次在植物中发现的基因组表观修饰现象,RdDM通过RNA-DNA序列相互作用直接导致DNA甲基化。植物中的RdDM和siRNA介导的mRNA降解现象,都是通过RNA使序列特异性基因发生沉默,它们对于植物的染色体重排、抵御病毒感染、基因表达调控和发育的许多过程起到了非常重要的作用。在植物中有很多的文献报道RdDM现象,但是对于其具体调控机理还不是很清楚。这里对RNA介导的植物DNA甲基化的基本特征进行了简要概述,主要对RdDM机理的研究进展进行了综述,其中包括RdDM过程中的DNA甲基转移酶的种类及其作用机理,DNA甲基化与染色质修饰之间的关系,以及与RdDM相关的重要蛋白质的研究等。在植物中,转录和转录后水平都可能发生RdDM,诱发基因沉默,前者常涉及靶基因启动子的甲基化,后者则牵涉到编码区的甲基化。RdDM的发生依赖于RNAi途径中相似的siRNA和酶,如DCL3、RdR2、SDE4和AGO4。植物中至少含有三类DNA甲基转移酶DRM1/2、MET1和CMT3,其作用部位是与RNA同源的DNA区域中的所有胞嘧啶,而组蛋白H3第九位赖氨酸的甲基化影响着胞嘧啶的甲基化。展开更多
Small RNAs are found in eukaryotes and are responsible for regulation of chromatin structure, RNA processing and stability, translation and transcription. 24-nt small interfering RNA (siRNA) are known to mediate gene ...Small RNAs are found in eukaryotes and are responsible for regulation of chromatin structure, RNA processing and stability, translation and transcription. 24-nt small interfering RNA (siRNA) are known to mediate gene inactivation via the RNA-directed DNA methylation pathway (RdDM) and are important for natural heritable changes in plant species. DNA cytosine methylation can be maintained between generations and this may be important for accelerated adaption to stress conditions. Research is currently focused toward the epigenetic response to disease, the stability of DNA methylation over generations, the elucidation of newly discovered pathways for de novo DNA methylation, and the application of epigenetic variation to breeding programs. This review aims to give a brief but comprehensive examination on small RNAs and transgenerational epigenetic variation.展开更多
It has been hypothesized that DNA damage has the potential to induce DNA hypermethylation,contributing to carcinogenesis in mammals.However,there is no sufficient evidence to support that DNA damage can cause genome-w...It has been hypothesized that DNA damage has the potential to induce DNA hypermethylation,contributing to carcinogenesis in mammals.However,there is no sufficient evidence to support that DNA damage can cause genome-wide DNA hypermethylation.In this study,we demonstrated that DNA single-strand breaks with 3′blocked ends(DNA 3′blocks)not only can reinforce DNA methylation at normally methylated loci but also can induce DNA methylation at normally nonmethylated loci in plants.The CG and CHG hypermethylation tend to localize within gene bodies,with a significant proportion being de novo generated.In contrast,the CHH hypermethylation is concentrated in centromeric and pericentromeric regions,primarily being reinforced methylation.Mechanistically,DNA 3′blocks regulate the DREAM complex to induce CG and CHG methylation.Moreover,they utilize the RdDM pathway to induce CHH hypermethylation.Intriguingly,repair of DNA damage or blocking the DNA damage response can fully abolish CHH hypermethylation and partially rescue CHG hypermethylation but rarely alter CG hypermethylation,indicating that DNA damage-induced symmetric DNA methylation can serve as a form of genetic imprinting.Collectively,these results suggest that DNA damage is an important force driving the emergence and evolution of genomic DNA methylation levels and patterns in plants.展开更多
DNA methylation and demethylation activities are coordinated to maintain DNA methylation patterns.However,the interplay between them remains to be explored.In this study,we found that loss of DNA demethylase in maize ...DNA methylation and demethylation activities are coordinated to maintain DNA methylation patterns.However,the interplay between them remains to be explored.In this study,we found that loss of DNA demethylase in maize results in a significant decrease in CHH methylation,a hallmark of RNA-directed DNA methylation(RdDM).Further analyses suggested that this is caused by reduced expression in the DNA demethylase mutant of three CLASSY(CLSY)genes,which encode chromatin remodelers controlling small RNA production.Series of molecular assays confirm that the expression of three maize CLSY genes is sensitive to the DNA methylation levels at their promoter regions,which are regulated by both the DNA demethylase and the RdDM pathway.Moreover,we revealed that Arabidopsis DNA demethylase mutants also show decreased CHH methylation and reduced expression of CLSY1,one of four CLSY genes in Arabidopsis.Similar to the observations in maize,the expression of CLSY1 is associated with DNA methylation levels of its promoter that is targeted by both DNA demethylase and RdDM pathways.Taken together,these results suggest a conserved interplay between DNA demethylation and RdDM pathways,revealing a mechanism to maintain the homeostasis of DNA methylation levels across plants.展开更多
The domains rearranged methyltransferases(DRMs)play a critical role in the RNA-directed DNA methylation(RdDM)pathway in plants.However,the effects of inactivating the RdDM pathway on gene expression,transposable eleme...The domains rearranged methyltransferases(DRMs)play a critical role in the RNA-directed DNA methylation(RdDM)pathway in plants.However,the effects of inactivating the RdDM pathway on gene expression,transposable element(TE)activity,and phenotype in soybean remain unexplored.Here,we employed clustered regularly interspaced short palindromic repeats(CRISPR)/CRISPR-associated protein 9 gene editing to generate a quintuple mutant line in soybean(Gmdrm2a^(−/−)2b^(−/−)2c^(−/−)3a^(−/−)3b^(−/−),designated Gmdrm).Gmdrm exhibited severe developmental abnormalities,including dwarfism and delayed growth,albeit remaining viable and fertile;however,the fully homozygous mutant could be maintained for a limited number of generations(T0–T3).Whole genome bisulfite sequencing revealed a significant reduction in DNA methylation across all cytosine sequence contexts,with an average loss of 10%.The loss of mC was biased toward euchromatic regions,which is in contrast to the chromomethylase mutant.Transcriptome profiling identified 1,685 up-regulated genes,including photosynthesis-related genes,accompanied with altered chloroplast ultrastructure.Additionally,a cluster of resistance(R)genes on chromosome 16 was significantly up-regulated,coinciding with their reduced non-CG methylation.We also observed 3,164 differentially expressed TEs(DETs),of which,2,655 were up-regulated and hypomethylated along their entire length.A substantial reduction in the abundance of 24-nt small interfering RNAs(siRNAs)in the Gmdrm mutant was detected by small RNA sequencing.Of note,the DRM-targeted TEs typically display higher levels of 24-nt siRNA abundance,shorter lengths,and are more AT-rich compared to chromomethylase-targeted TEs,highlighting 24-nt siRNAs as key determinants of DRM-dependent TE regulation.Together,this study documents a critical role of DRM-mediated DNA methylation in regulating gene expression,TE silencing,and normal development in soybean.展开更多
RNA-directed DNA methylation (RdDM) is a nuclear process in which small interfering RNAs (siRNAs) direct the cytosine methylation of DNA sequences that are complementary to the siRNAs. In plants, double stranded-RNAs ...RNA-directed DNA methylation (RdDM) is a nuclear process in which small interfering RNAs (siRNAs) direct the cytosine methylation of DNA sequences that are complementary to the siRNAs. In plants, double stranded-RNAs (dsRNAs) generated by RNA-dependent RNA polymerase 2 (RDR2) serve as precursors for Dicer-like 3 dependent biogenesis of 24-nt siRNAs. Plant specific RNA polymerase IV (Pol IV) is presumed to generate the initial RNA transcripts that are substrates for RDR2. siRNAs are loaded onto an argonaute4-containing RISC (RNA-induced silencing complex) that targets the de novo DNA methyltransferase DRM2 to RdDM target loci. Nascent RNA transcripts from the target loci are generated by another plant-specific RNA polymerase, Pol V, and these transcripts help recruit com- plementary siRNAs and the associated RdDM effector complex to the target loci in a transcrip- tion-coupled DNA methylation process. Small RNA binding proteins such as ROS3 may direct tar- get-specific DNA demethylation by the ROS1 family of DNA demethylases. Chromatin remodeling en- zymes and histone modifying enzymes also participate in DNA methylation and possibly demethylation. One of the well studied functions of RdDM is transposon silencing and genome stability. In addition, RdDM is important for paramutation, imprinting, gene regulation, and plant development. Lo- cus-specific DNA methylation and demethylation, and transposon activation under abiotic stresses suggest that RdDM is also important in stress responses of plants. Further studies will help illuminate the functions of RdDM in the dynamic control of epigenomes during development and environmental stress responses.展开更多
Tillering is a major determinant of rice plant architecture and grain yield.Here,we report that depletion of rice OsNRPD1a and OsNRPD1b,two orthologs of the largest subunit of RNA polymerase IV,leads to a high-tilleri...Tillering is a major determinant of rice plant architecture and grain yield.Here,we report that depletion of rice OsNRPD1a and OsNRPD1b,two orthologs of the largest subunit of RNA polymerase IV,leads to a high-tillering phenotype,in addition to dwarfism and smaller panicles.OsNRPD1a and OsNRPD1b are required for the production of 24-nt small interfering RNAs that direct DNA methylation at transposable elements(TEs)including miniature inverted-repeat TEs(MITEs).Interestingly,many genes are regulated either positively or negatively by TE methylation.Among them,OsMIR156d and OsMIR156j,which promote rice tillering,are repressed by CHH methylation at two MITEs in the promoters.By contrast,D14,which suppresses rice tillering,is activated by CHH methylation at an MITE in its downstream.Our findings reveal regulation of rice tillering by RNA-directed DNA methylation at MITEs and provide potential targets for agronomic trait enhancement through epigenome editing.展开更多
Epigenetic gene regulation involves the stable propagation of gene activity states through mitotic, and sometimes even meiotic, cell divisions without changes in DNA sequence. Paramutation is an epigenetic phenomenon ...Epigenetic gene regulation involves the stable propagation of gene activity states through mitotic, and sometimes even meiotic, cell divisions without changes in DNA sequence. Paramutation is an epigenetic phenomenon involving changes in gene expression that are stably transmitted through mitosis as well as meiosis. These heritable changes are mediated by in trans interactions between homologous DNA sequences on different chromosomes. During these in trans interactions, epigenetic information is transferred from one allele of a gene to another allele of the same gene, resulting in a change in gene expression. Although paramutation was initially discovered in plants, it has recently been observed in mammals as well, suggesting that the mechanisms underlying paramutation might be evolutionarily conserved. Recent findings point to a crucial role for small RNAs in the paramutation process. In mice, small RNAs appear sufficient to induce paramutation, whereas in maize, it seems not to be the only player in the process. In this review, potential mechanisms are discussed in relation to the various paramutation phenomena.展开更多
文摘RNA介导的DNA甲基化作用(RNA-directed DNA Methylation,RdDM)是首次在植物中发现的基因组表观修饰现象,RdDM通过RNA-DNA序列相互作用直接导致DNA甲基化。植物中的RdDM和siRNA介导的mRNA降解现象,都是通过RNA使序列特异性基因发生沉默,它们对于植物的染色体重排、抵御病毒感染、基因表达调控和发育的许多过程起到了非常重要的作用。在植物中有很多的文献报道RdDM现象,但是对于其具体调控机理还不是很清楚。这里对RNA介导的植物DNA甲基化的基本特征进行了简要概述,主要对RdDM机理的研究进展进行了综述,其中包括RdDM过程中的DNA甲基转移酶的种类及其作用机理,DNA甲基化与染色质修饰之间的关系,以及与RdDM相关的重要蛋白质的研究等。在植物中,转录和转录后水平都可能发生RdDM,诱发基因沉默,前者常涉及靶基因启动子的甲基化,后者则牵涉到编码区的甲基化。RdDM的发生依赖于RNAi途径中相似的siRNA和酶,如DCL3、RdR2、SDE4和AGO4。植物中至少含有三类DNA甲基转移酶DRM1/2、MET1和CMT3,其作用部位是与RNA同源的DNA区域中的所有胞嘧啶,而组蛋白H3第九位赖氨酸的甲基化影响着胞嘧啶的甲基化。
文摘Small RNAs are found in eukaryotes and are responsible for regulation of chromatin structure, RNA processing and stability, translation and transcription. 24-nt small interfering RNA (siRNA) are known to mediate gene inactivation via the RNA-directed DNA methylation pathway (RdDM) and are important for natural heritable changes in plant species. DNA cytosine methylation can be maintained between generations and this may be important for accelerated adaption to stress conditions. Research is currently focused toward the epigenetic response to disease, the stability of DNA methylation over generations, the elucidation of newly discovered pathways for de novo DNA methylation, and the application of epigenetic variation to breeding programs. This review aims to give a brief but comprehensive examination on small RNAs and transgenerational epigenetic variation.
基金supported by the National Natural Science Foundation of China(grant no.32270288 to W.Q.grant no.32400245 to J.L.)+1 种基金the China Postdoctoral Science Foundation(2024M760519 to W.L.)the Beijing Life Science Academy(2024500CA0010 to W.Q.).
文摘It has been hypothesized that DNA damage has the potential to induce DNA hypermethylation,contributing to carcinogenesis in mammals.However,there is no sufficient evidence to support that DNA damage can cause genome-wide DNA hypermethylation.In this study,we demonstrated that DNA single-strand breaks with 3′blocked ends(DNA 3′blocks)not only can reinforce DNA methylation at normally methylated loci but also can induce DNA methylation at normally nonmethylated loci in plants.The CG and CHG hypermethylation tend to localize within gene bodies,with a significant proportion being de novo generated.In contrast,the CHH hypermethylation is concentrated in centromeric and pericentromeric regions,primarily being reinforced methylation.Mechanistically,DNA 3′blocks regulate the DREAM complex to induce CG and CHG methylation.Moreover,they utilize the RdDM pathway to induce CHH hypermethylation.Intriguingly,repair of DNA damage or blocking the DNA damage response can fully abolish CHH hypermethylation and partially rescue CHG hypermethylation but rarely alter CG hypermethylation,indicating that DNA damage-induced symmetric DNA methylation can serve as a form of genetic imprinting.Collectively,these results suggest that DNA damage is an important force driving the emergence and evolution of genomic DNA methylation levels and patterns in plants.
基金supported by the Foundation for Innovative Research Groups of the National Natural Science Foundation of China(32321005)the 111 Project Crop Genomics and Molecular Breeding(B20051)+1 种基金the Outstanding Youth Team Cultivation Project of Central Universities(2662023PY007)the Postdoctoral Fellowship Program of CPSF undergrantnumberGZB20240254.
文摘DNA methylation and demethylation activities are coordinated to maintain DNA methylation patterns.However,the interplay between them remains to be explored.In this study,we found that loss of DNA demethylase in maize results in a significant decrease in CHH methylation,a hallmark of RNA-directed DNA methylation(RdDM).Further analyses suggested that this is caused by reduced expression in the DNA demethylase mutant of three CLASSY(CLSY)genes,which encode chromatin remodelers controlling small RNA production.Series of molecular assays confirm that the expression of three maize CLSY genes is sensitive to the DNA methylation levels at their promoter regions,which are regulated by both the DNA demethylase and the RdDM pathway.Moreover,we revealed that Arabidopsis DNA demethylase mutants also show decreased CHH methylation and reduced expression of CLSY1,one of four CLSY genes in Arabidopsis.Similar to the observations in maize,the expression of CLSY1 is associated with DNA methylation levels of its promoter that is targeted by both DNA demethylase and RdDM pathways.Taken together,these results suggest a conserved interplay between DNA demethylation and RdDM pathways,revealing a mechanism to maintain the homeostasis of DNA methylation levels across plants.
基金supported by the National Key Research and Development Program of China(2022YFF1003303,2022YFD1201400)the National Natural Science Foundation of China(32301796,32272064)+1 种基金the Fundamental Research Funds for the Central Universities(2412023QD021,2412023YQ005)the China Postdoctoral Science Foundation grant(2019M651236).
文摘The domains rearranged methyltransferases(DRMs)play a critical role in the RNA-directed DNA methylation(RdDM)pathway in plants.However,the effects of inactivating the RdDM pathway on gene expression,transposable element(TE)activity,and phenotype in soybean remain unexplored.Here,we employed clustered regularly interspaced short palindromic repeats(CRISPR)/CRISPR-associated protein 9 gene editing to generate a quintuple mutant line in soybean(Gmdrm2a^(−/−)2b^(−/−)2c^(−/−)3a^(−/−)3b^(−/−),designated Gmdrm).Gmdrm exhibited severe developmental abnormalities,including dwarfism and delayed growth,albeit remaining viable and fertile;however,the fully homozygous mutant could be maintained for a limited number of generations(T0–T3).Whole genome bisulfite sequencing revealed a significant reduction in DNA methylation across all cytosine sequence contexts,with an average loss of 10%.The loss of mC was biased toward euchromatic regions,which is in contrast to the chromomethylase mutant.Transcriptome profiling identified 1,685 up-regulated genes,including photosynthesis-related genes,accompanied with altered chloroplast ultrastructure.Additionally,a cluster of resistance(R)genes on chromosome 16 was significantly up-regulated,coinciding with their reduced non-CG methylation.We also observed 3,164 differentially expressed TEs(DETs),of which,2,655 were up-regulated and hypomethylated along their entire length.A substantial reduction in the abundance of 24-nt small interfering RNAs(siRNAs)in the Gmdrm mutant was detected by small RNA sequencing.Of note,the DRM-targeted TEs typically display higher levels of 24-nt siRNA abundance,shorter lengths,and are more AT-rich compared to chromomethylase-targeted TEs,highlighting 24-nt siRNAs as key determinants of DRM-dependent TE regulation.Together,this study documents a critical role of DRM-mediated DNA methylation in regulating gene expression,TE silencing,and normal development in soybean.
基金supported by National Institutes of Health Grants R01GM070795 and R01GM059138. V.C. is thankful to ICAR, New Delhi for providing study leave.
文摘RNA-directed DNA methylation (RdDM) is a nuclear process in which small interfering RNAs (siRNAs) direct the cytosine methylation of DNA sequences that are complementary to the siRNAs. In plants, double stranded-RNAs (dsRNAs) generated by RNA-dependent RNA polymerase 2 (RDR2) serve as precursors for Dicer-like 3 dependent biogenesis of 24-nt siRNAs. Plant specific RNA polymerase IV (Pol IV) is presumed to generate the initial RNA transcripts that are substrates for RDR2. siRNAs are loaded onto an argonaute4-containing RISC (RNA-induced silencing complex) that targets the de novo DNA methyltransferase DRM2 to RdDM target loci. Nascent RNA transcripts from the target loci are generated by another plant-specific RNA polymerase, Pol V, and these transcripts help recruit com- plementary siRNAs and the associated RdDM effector complex to the target loci in a transcrip- tion-coupled DNA methylation process. Small RNA binding proteins such as ROS3 may direct tar- get-specific DNA demethylation by the ROS1 family of DNA demethylases. Chromatin remodeling en- zymes and histone modifying enzymes also participate in DNA methylation and possibly demethylation. One of the well studied functions of RdDM is transposon silencing and genome stability. In addition, RdDM is important for paramutation, imprinting, gene regulation, and plant development. Lo- cus-specific DNA methylation and demethylation, and transposon activation under abiotic stresses suggest that RdDM is also important in stress responses of plants. Further studies will help illuminate the functions of RdDM in the dynamic control of epigenomes during development and environmental stress responses.
基金This work was supported by grants from National Natural Science Foundation of China(grant no.31788103)to J.L.and Y.Q.and the National Key R&D Program of China(grant no.2016YFA0500800)to Y.Q.Y.Q.is a visiting investigator of the CAS Center for Excellence in Molecular Plant Sciences.
文摘Tillering is a major determinant of rice plant architecture and grain yield.Here,we report that depletion of rice OsNRPD1a and OsNRPD1b,two orthologs of the largest subunit of RNA polymerase IV,leads to a high-tillering phenotype,in addition to dwarfism and smaller panicles.OsNRPD1a and OsNRPD1b are required for the production of 24-nt small interfering RNAs that direct DNA methylation at transposable elements(TEs)including miniature inverted-repeat TEs(MITEs).Interestingly,many genes are regulated either positively or negatively by TE methylation.Among them,OsMIR156d and OsMIR156j,which promote rice tillering,are repressed by CHH methylation at two MITEs in the promoters.By contrast,D14,which suppresses rice tillering,is activated by CHH methylation at an MITE in its downstream.Our findings reveal regulation of rice tillering by RNA-directed DNA methylation at MITEs and provide potential targets for agronomic trait enhancement through epigenome editing.
文摘Epigenetic gene regulation involves the stable propagation of gene activity states through mitotic, and sometimes even meiotic, cell divisions without changes in DNA sequence. Paramutation is an epigenetic phenomenon involving changes in gene expression that are stably transmitted through mitosis as well as meiosis. These heritable changes are mediated by in trans interactions between homologous DNA sequences on different chromosomes. During these in trans interactions, epigenetic information is transferred from one allele of a gene to another allele of the same gene, resulting in a change in gene expression. Although paramutation was initially discovered in plants, it has recently been observed in mammals as well, suggesting that the mechanisms underlying paramutation might be evolutionarily conserved. Recent findings point to a crucial role for small RNAs in the paramutation process. In mice, small RNAs appear sufficient to induce paramutation, whereas in maize, it seems not to be the only player in the process. In this review, potential mechanisms are discussed in relation to the various paramutation phenomena.
