DNA MINOR GROOVE BINDING POLYAMIDES

描述(逐字摘自申请者摘要)：这项研究计划 寻求设计新的策略来合理地抑制介导的基因表达 转录因子(TF)。该策略是设计模块化的聚酰胺DNA 有效和特异地抑制转铁蛋白与基因结合的次要结合剂 启动子和干扰基因的表达。为此目的，一名 Ets致癌蛋白家族ELK-1与血清反应元件结合 选择c-fos启动子中的(SRE)作为模型转录因子。 对ELK-1与SRE结合的干扰将是定量的 通过迁移率变化分析来测量，以评估聚酰胺的抑制能力 Tf/DNA复合体。将使用定量的足迹分析来确定 其中聚酰胺结合在SRE启动子元件上，也结合在SRE启动子元件上 有约束力的。高效、特异的ELK-1/SRE抑制剂--聚酰胺 复杂的形成将被评估它们抑制的能力 C-fos启动子在无细胞和细胞内的转录激活 化验。有希望的代理人将被评估他们的干扰能力 一种与c-fos基因表达相关的肿瘤表型。同时， 聚酰胺将在保持目标的同时优化细胞活性 专一性。具体目标：目标1.聚酰胺DNA小沟槽的鉴定 干扰ELK-1与c-fos启动子结合的结合剂 在无细胞条件下。AIM2.聚酰胺DNA小槽结合效果的研究 ELK-1介导的c-fos启动子在无细胞条件下基因表达的研究 条件。Aim3.聚酰胺DNA小槽结合剂对ELK-1的影响 C-fos启动子在完整细胞中的基因表达。

DESCRIPTION (verbatim from the applicant's abstract): This research program seeks to devise new strategies to rationally inhibit gene expression mediated by transcription factors (TFs). The strategy is to design modular polyamide DNA minor binding agents that potently and specifically inhibit TF binding to gene promoters and disrupt gene expression. For this purpose, a representative of the Ets oncogenic protein family Elk-1, which binds a serum response element (SRE) contained within the c-fos promoter, was chosen as a model TF. Interference with the binding of Elk-1 to the SRE will be quantitatively measured by mobility shift assays, to assess a polyamide's ability to inhibit TF/DNA complexes. Quantitative footprint analysis will be used to determine where polyamides bind on the SRE promoter element and also the strength of the binding. Polyamides that are potent and specific inhibitors of Elk-1/SRE complex formation will be evaluated for their ability to inhibit transcriptional activation from the c-fos promoter in cell-free and cellular assays. Promising agents will be evaluated for their ability to interfere with a neoplastic phenotype associated with c-fos gene expression. In parallel, polyamides will be optimized for cellular activity while maintaining target specificity. Specific aims: Aim1. Identification of polyamide DNA minor groove binding agents that interfere with the binding of Elk-1 to the c-fos promoter under cell-free conditions. Aim2. Effects of polyamide DNA minor groove binding agents on Elk-1 mediated gene expression of the c-fos promoter under cell-free conditions. Aim3. Effects of polyamide DNA minor groove binding agents on Elk-1 mediated gene expression of the c-fos promoter in intact cells.