ANALYSIS OF AN OLIGO(DA) OLIGO(DT) BINDING PROTEIN

描述(改编自申请人的摘要)：关键控制点 在转录中可以由DNA结合蛋白控制，识别 基因组中的少量靶标。对这些具体情况的研究 交易因素导致了对许多 细胞过程，包括那些控制细胞周期进程的过程，以及 正常和异常的生长和分化。我们所知的相对较少 关于蛋白质与高度重复的DNA元件的特定相互作用 以及这些相互作用可能产生的转录影响。 简单非交替序列dA.dT在许多情况下高度重复 有机体。这一序列已经被证明采用了一种不同寻常的 构象，促进酵母中的转录。有人提议， 这种转录激活与其独特的结构有关。 DA.dt.调查人员最近报告了一种新的酵母的发现 与非交替dA.dT特异相互作用的DNA结合蛋白。 该蛋白(DAT1)已被纯化，并克隆了它的基因(DAT1)。更多 最近的研究表明，Datin可以负向调节 含有上游dA.dT区的基因。此外，达廷还能够 正向调节自身表达(Datin的启动子包含dA.dT 序列)。由于许多酵母启动子包含dA.dT区段，因此这些 观察表明，Datin可以调节大分子的表达 基因的数量。 这是这项提案中描述的两个主要目标。首先， 研究人员将确定达汀结合的分子基础 DA.dt.具体地说，调查员将定义不同形式的 Datin存在于体内，这些不同的精确DNA元素 Datin的形式与之相互作用，以及Datin中的氨基酸残基 是这种互动所必需的。调查员还将试图 产生用于X-射线衍射研究的Datin/寡聚(Da)(DT)晶体。 其次，调查者将确定什么是生理功能 达丁是。具体地说，调查员将确定 Datin对含有以下成分的基因的转录调控作用 上游dA.dT区域，并识别和表征其他基因 对与DAT1函数重叠的函数进行编码。这些研究将不会 仅有助于理解分子机制 负责特定的DNA/蛋白质相互作用，但也将做出贡献 对基因表达调控的理解。

DESCRIPTION (Adapted from applicant's abstract): Critical control points in transcription can be governed by DNA binding proteins that recognize small numbers of targets within the genome. Studies of these specific trans-acting factors have led to insights into the regulation of many cellular processes including those that govern cell cycle progression, and normal and aberrant growth and differentiation. Relatively little is known about specific interactions of proteins with highly repeated DNA elements and the transcriptional influences that these interactions can exert. The simple nonalternating sequence dA.dT is highly repeated in many organisms. This sequence, which has been shown to adopt an unusual conformation, promotes transcription in yeast. It has been proposed that this transcriptional activation is related to the unique structure of dA.dT. The investigator recently reported the discovery of a new yeast DNA-binding protein that specifically interacts with nonalternating dA.dT. This protein (datin) has been purified, and its gene, (DAT1) cloned. More recent work shows that datin can negatively regulate the expression of genes which contain upstream dA.dT tracts. In addition, datin is able to positively regulate its own expression (datin's promoter contains dA.dT sequences). Since many yeast promoters contain dA.dT tracts, these observations suggest that datin can modulate the expression of large numbers of genes. These are two major objectives described in this proposal. First, the investigator will determine the molecular basis by which datin binds to dA.dT. Specifically the investigator will define the different forms of datin which exist in vivo, the precise DNA elements that these different forms of datin interact with, and the amino acid residues in datin which are necessary for this interaction. The investigator will also attempt to generate datin/oligo(dA).(dT) crystals for x-ray diffraction studies. Second, the investgator will determine what the physiological functions of datin are. Specifically, the investigator will determine the transcriptional regulatory effects that datin exerts on genes which contain upstream dA.dT tracts, and identify and characterize additional genes which encode functions that overlap the function of DAT1. These studies will not only contribute to the understanding of the molecular mechanisms responsible for specific DNA/protein interactions, but will also contribute to the understanding of the regulation of gene expression.