Dynamic 31P NMR of Backbone Dynamics in DNA

描述（申请人提供）：表观遗传学是基于基因表达的信息遗传。它决定了何时何地使用遗传信息。人类最常见的表观遗传修饰是DNA序列CpG步骤中胞嘧啶的甲基化（Jones and Baylin 2002）。这些步骤的甲基化状态在印迹和基因沉默中起着至关重要的作用。这一点非常重要，因为甲基化诱导的基因沉默被认为是许多人类癌症的前兆（Jones and Laird 1999）。虽然已经做了很多工作来了解CpG甲基化在肿瘤发生中的作用，但甲基化的结构和动力学效应尚未得到很好的理解。

DESCRIPTION (provided by applicant): Epigenetics is the inheritance of information based on gene expression. It determines when and where genetic information will be used. The most common human epigenetic modification is the methylation of cytosines in CpG steps of DNA sequences (Jones and Baylin 2002). The methylation state of these steps is known to play a crucial role in imprinting and gene silencing. This is tremendously important as methylation induced gene silencing is considered a precursor in many human cancers (Jones and Laird 1999). While much work has been done to understand the role of CpG methylation in tumorgenesis, the structural and dynamical effects of methylation are not well understood. Our research focuses on how proteins recognize specific DNA sequences and the effects of cytosine methylation on this recognition. In particular, we study the role of DNA dynamics in this recognition process. Our previous solid-state deuterium NMR studies of methylated and unmethylated DNA binding sites have shown that cytosine methylation quenches DNA backbone dynamics (Geahigan, Meints et al. 2000; Meints and Drobny 2001). However, these experiments required extensive sample preparation and allowed the study of only one nucleotide step at a time. We propose to use phosphorus NMR and complete lineshape analysis to investigate the backbone dynamics of each step in methylated and unmethylated DNA samples. This will allow us to rapidly probe a myriad of DNA binding sites for dynamical effects of cytosine methylation. The ability to quickly study a large number of binding sites will uncover any trends in methylation effects, which we hope can then be used to design tools for early detection of cancers. Oligonucleotide Sequences for 31P Dynamic NMR Study A. The EcoRI binding site, d(CGCGAATTCGCG)2. B. The CRE containing sites, d(ATGACGTCAT)2 and d(GAAAACGTTTTC)2. C. The cMyc binding site, d(ACCACGTGGT)2.