Epigenetic Variation in Mice with Defective Imprinting

描述（由申请人提供）：基因组印记是一种分子过程，通过不同的表观遗传标记来区分基因的母系和父系等位基因。这个过程的核心是差异甲基化结构域（dmd）的产生，其中一个亲本等位基因被甲基化，而另一个等位基因未甲基化。dmd通过两步机制发展，其中DNA胞嘧啶甲基化模式（印记）首先被放置在亲本等位基因之一上（取决于基因在卵母细胞或精子中），然后这种表观遗传修饰被维持（遗传）。dmd控制着发育中的胎儿和相关胚胎外组织中大部分（如果不是全部的话）印迹基因的印记。我们的研究结果表明，在胚胎着床前发育过程中，DMD序列甲基化标记的维持对印迹过程至关重要，从而对胚胎发育至关重要。本研究的主要目的是研究DNMT1 DNA胞嘧啶甲基转移酶dnmt10和DNMT1在这一维持过程中的作用。DNMT1在体细胞中表达，dnmt10是一种源自卵母细胞的DNMT1变体，在卵母细胞中表达，也在着床前发育期间表达。这些研究分为两个目标，每个目标都针对一个特定的假设：(1)我们假设在着床前发育过程中基因组印记的遗传取决于卵母细胞和受精卵来源的DNMT1蛋白的维持甲基化活性。关于植入前dnmt10和Dnmt1作用的假设将在多种实验中得到验证，这些实验将检验Dnmt1基因突变对小鼠胚胎干细胞基因组印迹维持的影响。(2)我们假设dnmt10和DNMT1蛋白的特定区域参与了基因组印迹的遗传，并且可以通过在体内研究合适的DNMT1突变体来推断这些区域的特定功能。

DESCRIPTION (provided by applicant): Genomic imprinting is a molecular process that distinguishes, with different epigenetic marks, the maternal and paternal alleles of genes. Central to this process is the creation of differentially methylated domains (DMDs) in which one parental allele is methylated and the opposite allele is unmethylated. DMDs develop through a two- step mechanism in which a DNA cytosine methylation pattern (imprint) is first placed on one of the parental alleles (in oocyte or sperm depending on the gene) and this epigenetic modification is then maintained (inherited). DMDs govern the imprinting of most, if not all, of the imprinted genes in the developing fetus and associated extraembryonic tissues. Our results indicate that the maintenance of methylation marks on DMD sequences during preimplantation development is crucial to the imprinting process and thus to the developing embryo. The main objective of this proposal is to study the roles of the DNMT1 DNA cytosine methyltransferases, DNMT1o and DNMT1s, in this maintenance process. DNMT1s is expressed in somatic cells and DNMT1o is an oocyte-derived DNMT1 variant that is expressed in oocytes and also during preimplantation development. The studies are divided into two Aims, each addressing a specific hypothesis: (1) We hypothesize here that the inheritance of genomic imprints during preimplantation development depends on maintenance methylation activities of both oocyte- and zygote-derived DNMT1 proteins. Hypotheses regarding the preimplantation roles of DNMT1o and DNMT1s will be tested in a variety of experiments examining the effect of Dnmt1 gene mutations on the maintenance of genomic imprints in mouse ES cells. (2) We hypothesize here that specific regions of the DNMT1o and DNMT1s proteins are involved in the inheritance of genomic imprints, and that the specific functions of these regions can be deduced by studying appropriate DNMT1 mutants in vivo.