The de novo cytosine methyltransferase DRM2 requires intact UBA domains and a catalytically mutated paralog DRM3 during RNA-directed DNA methylation in Arabidopsis thaliana.
The de novo cytosine methyltransferase DRM2 requires intact UBA domains and a catalytically mutated paralog DRM3 during RNA-directed DNA methylation in Arabidopsis thaliana.
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DOI：
10.1371/journal.pgen.1001182
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发表时间：
2010-10-28
期刊：
影响因子：
4.5
通讯作者：
Jacobsen SE
中科院分区：
文献类型：
--
作者：
Henderson IR;Deleris A;Wong W;Zhong X;Chin HG;Horwitz GA;Kelly KA;Pradhan S;Jacobsen SE
真核DNA胞嘧啶甲基化可用于转录沉默重复序列，包括转座子和逆转录病毒。这种沉默在细胞世代之间是稳定的，因为胞嘧啶甲基化通过DNA复制在表观遗传学上得以维持。拟南芥Dnmt 3胞嘧啶甲基转移酶直系同源物重排甲基转移酶2（DRM 2）是建立小干扰RNA（siRNA）指导的DNA甲基化所必需的。在哺乳动物中，PIWI蛋白和皮尔纳在收敛进化的RNA指导的DNA甲基化系统中起作用，该系统是抑制生殖系中转座子表达所需的。从头甲基化也可能独立于RNA干扰和小RNA，如粗糙脉孢菌。在这里，我们确定了催化突变的DRM 2旁系同源开花植物基因组，在拟南芥中，我们称之为结构域重排甲基转移酶3（DRM 3）。尽管被催化突变，但DRM 3是正常维持非CG DNA甲基化、建立由重复序列触发的RNA指导的DNA甲基化和积累重复相关小RNA所必需的。虽然哺乳动物无催化活性的Dnmt 3L旁系同源物以类似的方式起作用，但系统发育分析表明DRM和Dnmt 3蛋白家族在植物和动物中独立地分化。我们还通过定点突变表明，DRM 2 N-末端乌巴结构域和C-末端甲基转移酶结构域都是正常RNA指导的DNA甲基化所必需的，支持乌巴结构域的基本靶向功能。这些结果表明，植物和哺乳动物的RNA指导的DNA甲基化系统的组合的祖先和收敛的功能。核DNA数量在物种之间变化很大，与基因数量相关性很差。基因组大小的变化可以用重复DNA的不同数量来解释。重复DNA可能是移动的，这意味着它可以增加其在基因组中的拷贝数。为了防止这种情况，植物和动物抑制重复序列的表达，通常通过在胞嘧啶碱基上用甲基标记重复序列。DRM 2是一种能够建立这种甲基化的酶，它可以通过短的互补RNA向导被引导到靶序列。由于重复序列倾向于产生短RNA，因此它们被有效地识别和沉默。我们发现，DRM 2需要一个相关的非活性DRM 3蛋白正常沉默重复序列。类似的情况也存在于哺乳动物中，其中活性和非活性DNA甲基转移酶共同作用以沉默重复序列。我们还表明，DRM 2酶的非催化区域在功能上是重要的，我们推测参与靶向酶的基因组。虽然植物和哺乳动物的RNA指导的DNA甲基化系统有着关键的相似之处，但也有重要的机制差异，这意味着它们可能是趋同出现的。
Eukaryotic DNA cytosine methylation can be used to transcriptionally silence repetitive sequences, including transposons and retroviruses. This silencing is stable between cell generations as cytosine methylation is maintained epigenetically through DNA replication. The Arabidopsis thaliana Dnmt3 cytosine methyltransferase ortholog DOMAINS REARRANGED METHYLTRANSFERASE2 (DRM2) is required for establishment of small interfering RNA (siRNA) directed DNA methylation. In mammals PIWI proteins and piRNA act in a convergently evolved RNA–directed DNA methylation system that is required to repress transposon expression in the germ line. De novo methylation may also be independent of RNA interference and small RNAs, as in Neurospora crassa. Here we identify a clade of catalytically mutated DRM2 paralogs in flowering plant genomes, which in A.thaliana we term DOMAINS REARRANGED METHYLTRANSFERASE3 (DRM3). Despite being catalytically mutated, DRM3 is required for normal maintenance of non-CG DNA methylation, establishment of RNA–directed DNA methylation triggered by repeat sequences and accumulation of repeat-associated small RNAs. Although the mammalian catalytically inactive Dnmt3L paralogs act in an analogous manner, phylogenetic analysis indicates that the DRM and Dnmt3 protein families diverged independently in plants and animals. We also show by site-directed mutagenesis that both the DRM2 N-terminal UBA domains and C-terminal methyltransferase domain are required for normal RNA–directed DNA methylation, supporting an essential targeting function for the UBA domains. These results suggest that plant and mammalian RNA–directed DNA methylation systems consist of a combination of ancestral and convergent features. Nuclear DNA quantity varies widely between species and is poorly correlated with gene number. Variation in genome size can be explained by differing amounts of repetitive DNA. Repetitive DNA may be mobile, meaning it can increase its copy number within genomes. To prevent this, plants and animals suppress expression of repeats, often by marking the repeated sequence with a methyl group on cytosine bases. DRM2 is an enzyme capable of establishing this methylation, which can be guided to target sequences by short complementary RNA guides. As repeated sequences are prone to generate short RNAs they are efficiently recognized and silenced. We show that DRM2 requires a related inactive DRM3 protein to normally silence repeated sequences. A similar situation exists in mammals, where active and inactive DNA methyltransferases act together to silence repeats. We also demonstrate that non-catalytic regions of the DRM2 enzyme are functionally important, which we speculate to be involved in targeting the enzyme to the genome. Although plant and mammal RNA–directed DNA methylation systems share key similarities, there are important mechanistic differences, meaning that they are likely to have arisen convergently.
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DOI：
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2000-04-25
影响因子：
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通讯作者：
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