DNA METHYLATION AND GENE SILENCING IN ARABIDOPSIS

胞嘧啶甲基化是在包括哺乳动物在内的真核生物基因组中发现的一种DNA修饰。植物和丝状真菌。甲基化与基因表达缺失（沉默）和同源重组抑制有关。维持正确的甲基化模式对发育至关重要。例如，在人类中，某些基因组区域甲基化模式的改变导致许多遗传疾病，包括智力迟钝综合征和易患恶性肿瘤。在小鼠中，胞嘧啶甲基转移酶敲除突变导致胚胎死亡。在高等植物模式生物拟南芥中，甲基化缺陷突变体表现出许多异常形态，但仍然具有活力和可育性。因此，拟南芥提供了一个非常容易处理的系统，可以通过分子和遗传方法研究甲基化和沉默的机制。我们的研究重点是拟南芥中一个甲基化内源基因家族，该家族由两个单线基因和三个非连锁基因组位置的尾对尾反向重复基因重复组成。先前的研究表明，反向重复位点触发了基因组中其他地方相同序列的甲基化。矛盾的是，虽然甲基化导致单线位点沉默，但甲基化的反向重复序列中的一个基因被表达。本建议的目的是了解倒置重复位点逃避甲基化通常抑制作用的机制。这个问题将通过描述染色质结构、顺式作用的DNA序列和控制沉默单线位点与表达的反向重复位点的甲基化和表达的反式作用因子来探讨。

Cytosine methylation is a DNA modification found in the genomes of eukaryotes including mammals, higher. plants, and filamentous fungi. Methylation is associated with loss of gene expression (silencing) and suppression of homologous recombination. Maintenance of correct methylation patterns is critically important for development. For example, in humans, alteration of methylation patterns in certain genomic regions leads to a number of genetic diseases including mental retardation syndromes and pre-disposition to malignancies. In mice, a cytosine methyltransferase knock-out mutation results in embryo-lethality. In the higher plant model organism Arabidopsis thaliana, methylation-deficient mutants display a number of aberrant morphologies but are nonetheless viable and fertile. Therefore, Arabidopsis provides an extremely tractable system to study the mechanisms of methylation and silencing with molecular and genetic approaches. Our research focuses on a family of methylated endogenous genes in Arabidopsis that consists of two singlet genes plus a tail-to-tail inverted repeat gene duplication at three unlinked genomic positions. Previous work showed that the inverted repeat locus triggers methylation of the identical sequences elsewhere in the genome. Paradoxically, although methylation causes silencing of a singlet locus, a gene in the methylated inverted repeat is expressed. The goal of this proposal is to understand the mechanisms by which the inverted repeat locus escapes the normally repressive effects of methylation. This question will be explored by characterizing the chromatin structure, cis-acting DNA sequences, and trans-acting factors that control methylation and expression of the silenced singlet locus versus the expressed inverted repeat locus.