Enhancement of Cellular DNA Flexibility

描述(申请人提供)：DNA是一种局部不可弯曲的聚合物。尽管如此，DNA固有的不灵活在细胞中被以某种方式克服了，允许DNA不断地折叠和循环以存储和基因表达。了解DNA在健康和患病细胞中的灵活性是如何增强的，这一点至关重要。HMGB蛋白是真核细胞染色质中含量丰富的非组蛋白。HMGB蛋白被认为是促进DNA弯曲和扭曲的结构因子。我们试图更好地理解HMGB蛋白的分子机制，以及HMGB蛋白在活细胞中基因表达中的功能作用。提出了四个具体目标。1.单分子实验表征HMGB机理。我们将测试HMGB蛋白是否会导致DNA中的瞬时柔性铰链。2.利用阳离子蛋白结构域分析DNA折叠。我们将测试HMGB蛋白是否可以使用阳离子结构域来导致DNA崩溃。3.细菌DNA易化环的分析。我们将测试HMGB蛋白是否可以稳定细菌抑制环，并取代细菌结构DNA结合蛋白。4.分析酵母中HMGB蛋白对转录激活子位置效应的影响。我们将测试HMGB蛋白是否稳定了酵母中的DNA调控环。LAV描述：构建生物的指令被编码在细胞食谱(DNA)中的食谱(基因)中。很长的DNA分子很难急剧弯曲和扭曲，但必须折叠成多种形状才能在细胞中发挥作用。通过了解使DNA变得灵活的细胞蛋白质，我们将了解可能用于人工调节疾病基因的工具。

DESCRIPTION (provided by applicant): DNA is a locally inflexible polymer. Nonetheless, the intrinsic inflexibility of DNA is somehow overcome in cells, allowing the constant folding and looping of DNA for storage and gene expression. It is fundamentally important to understand how DNA flexibility is enhanced in healthy, and diseased cells. HMGB proteins are abundant non-histone proteins in eukaryotic chromatin. HMGB proteins are thought to function as architectural factors that enhance DNA bending and twisting. We seek to better understand, both the molecular mechanism of HMGB proteins, and the functional role of HMGB proteins in gene expression in living cells. Four specific aims are proposed. 1. Characterization of HMGB mechanism by single molecule experiments. We will test if HMGB proteins cause transient flexible hinges in DNA. 2. Analysis of DNA collapse by cationic protein domains. We will test if HMGB proteins can use cationic domains to cause DNA collapse. 3. Analysis of facilitated DNA looping in bacteria. We will test if HMGB proteins can stabilize bacterial repression loops and substitute for bacterial architectural DNA binding proteins. 4. Analysis of the effect of HMGB proteins on transcription activator position effects in yeast. We will test if HMGB proteins stabilize DNA regulatory loops in yeast. Lav description: The instructions for building living things are coded in recipes (genes) within the cell cookbook (DNA). Very long DNA molecules are difficult to sharply bend and twist, but must nonetheless be folded into many shapes in order to function in cells. By understanding the cell proteins that make DNA flexible, we will learn about tools that may be used to artificially regulate disease genes.