Genetics of DNA Methylation Patterning of Arabidopsis

描述（由申请人提供）：胞嘧啶DMA甲基化是基因沉默的表观遗传标记，在许多基因调控系统中很重要，包括基因组印记、x染色体失活、转座子和其他含有直接或反向重复序列的DNA序列的沉默。甲基化在癌症生物学中很重要，因为肿瘤经常表现出特定肿瘤抑制基因的全基因组去甲基化和高甲基化。我们正在研究模式植物拟南芥DNA甲基化控制的机制。拟南芥DNA甲基化系统与哺乳动物系统有很多共同之处，显示出三种人类DNA甲基转移酶Dnmtl， 2和3的同源物。正遗传学和反向遗传学都可以进行，拟南芥可以忍受几乎消除甲基化的突变，从而可以进行详细的分析。我们最近发现，包括DICER、RNA依赖性RNA聚合酶和ARGONAUTE蛋白在内的RNA沉默组分是DNA甲基化建立和非cg环境下甲基化维持机制的核心。我们提出了一系列的遗传和生化实验来进一步研究这一现象。直接重复序列和反向重复序列可以通过不同的机制触发DNA从头甲基化，我们建议研究这些差异。我们还提出了旨在分离这些过程中重要因素的突变筛选。ARGONAUTE4蛋白可能在小rna和染色质修饰酶的界面上起作用，我们拟详细研究该蛋白的作用机制。由于DNA甲基化控制机制的许多方面在不同的真核生物中是相似的，因此在拟南芥中取得的进展很可能会揭示一系列涉及基因组防御、发育生物学和人类疾病的表观遗传调控系统。

DESCRIPTION (provided by applicant): Cytosine DMA methylation is an epigenetic mark for gene silencing that is important in many gene regulatory systems including genomic imprinting, X-chromosome inactivation, and the silencing of transposons and other DNA sequences containing either direct or inverted repeats. Methylation is important in cancer biology, as tumors often show both genome wide demethylation and hypermethylation of specific tumor suppressor genes. We are studying the mechanisms of DNA methylation control in the model plant Arabidopsis thaliana. Arabidopsis DNA methylation systems have much in common with mammalian systems, showing homologs of the three human DNA methyltransferases, Dnmtl, 2, and 3. Both forward and reverse genetics can be performed, and Arabidopsis can tolerate mutations that virtually eliminate methylation, allowing for detailed analysis. We have recently discovered that RNA silencing components including a DICER, an RNA-dependent RNA polymerase, and an ARGONAUTE protein, are central to the mechanisms by which DNA methylation is established, and by which methylation in non-CG contexts is maintained. We propose a series of genetic and biochemical experiments to further study this phenomena. Direct repeats and inverted repeats can trigger de novo DNA methylation by different mechanisms, and we propose to study these differences. We also propose mutant screens aimed at isolating factors important in these processes. The ARGONAUTE4 protein likely acts at the interface between small RNAs and chromatin modifying enzymes, and we propose to study the mechanism of action of this protein in detail. Since many aspects of DNA methylation control mechanisms are similar in diverse eukaryotic organisms, it is likely that progress made in Arabidopsis will shed light on a wide array of epigenetic regulatory systems involved in genome defense, developmental biology, and human disease.