PEPTIDE NUCLEIC ACIDS (PNA)-MEDIATED GENE EXPRESSION

描述：(摘自申请者的摘要)许多疾病都是由 导致基因产物缺陷的突变或由于基因 对基因表达的调节，导致不适当的合成。人类 细胞包含许多与结构或功能相关的基因，它们可以补偿 突变基因产物的功能。不幸的是，这些基因中的大多数是 要么发育受调控，要么在特定组织中表达，要么受调控 在上游组件的路径中，其功能可能由于以下原因而丢失 基因的突变。 这个项目的最终目标是诱导这些 并将其应用于人类疾病的基因治疗 新技术，肽核酸(PNAS)诱导基因表达。PNAS 是带有修饰骨架的合成寡核苷酸。PNA可以绑定到 PNA结合部位为双链结构。这种D-LOOP结构具有 被发现具有启动基因转录的能力。在私家侦探的 以前的研究表明，PNA与伽马珠蛋白基因中的一个目标位置的结合 发现可诱导绿色荧光蛋白报告基因和内源性丙种球蛋白基因 表情。然而，PNA结合诱导基因的分子机制 表达还没有被研究过。影响PNA介导的内分泌的参数 活体基因的表达也没有被研究过。 在这项提案中，首席研究员计划研究分子 PNA结合诱导基因表达的机制。D-环路长度 将定义基因转录启动的要求。这个 参与PNA结合诱导基因表达的转录成分将 要下定决心。来自PNA启动子的RNA转录本的结构将 被研究。PNA结合介导的基因表达与基因表达的相关性 将对DNA修复/复制进行调查。体内影响因素的研究 PNA基因的诱导，如PNA递送效率和细胞内PNA 绑定也将被研究。从这些研究中获得的知识将 有助于理解PNA结合诱导基因的分子基础 表情。从这些研究中获得的知识也将用于 探索利用PNA技术诱导两种基因表达的可能性 治疗人类两种最常见疾病的人类基因：诱导 镰状细胞病的γ-珠蛋白基因表达及p21的诱导 癌症的基因表达。从这个项目中获得的知识也将 有助于理解PNA结合诱导基因表达的分子机制 并为治疗许多人类疾病提供直接证据，包括 遗传病和癌症。目标是： 探讨PNA诱导基因表达的分子机制。 研究影响PNA递送效率和细胞内的参数 PNA结合。 探索PNA结合诱导内源性人类基因的可能性 表情。

DESCRIPTION: (from the applicant's abstract) Many diseases are caused by mutations that lead to defective gene products or by disturbance in the regulation of gene expression that results in inappropriate synthesis. Human cells contain many structure- or function-related genes that can compensate the function of the mutant gene product. Unfortunately, most of these genes are either developmentally regulated, expressed in specific tissues, or regulated in a pathway by an upstream component whose function may be lost due to mutations in the gene. The ultimate goal of this project is to induce the expression of these function-related genes and apply it to gene therapy of human diseases using a novel technology, peptide nucleic acids (PNAs)-induced gene expression. PNAs are synthetic oligonucleotides with modified backbones. PNAs can bind to double-stranded structures at the PNA binding site. This D-loop structure has been found to have the capability to initiate gene transcription. In the PI's previous studies, binding of PNAs to a target site in the gamma-globin gene was found to induce both the GFP reporter gene and endogenous gamma-globin gene expression. However, the molecular mechanisms of PNA binding-induced gene expression have not been studied. The parameters that affect PNA-mediated in vivo gene expression also have not been investigated. In this proposal the principal investigator plans to study the molecular mechanisms of PNA binding-induced gene expression. The D-loop length requirement for gene transcription initiation will be defined. The transcription components involved in PNA binding-induced gene expression will be determined. The structure of the RNA transcripts from the PNA promoter will be studied. The correlation between PNA binding-mediated gene expression and DNA repair/replication will be investigated. The parameters affecting in vivo PNA gene induction such as PNA delivery efficiency and intracellular PNA binding also will be studied. The knowledge obtained from these studies will contribute to understanding the molecular basis of PNA binding-induced gene expression. The knowledge obtained from these studies will also be used to explore the possibility of using PNA technology to induce the expression of two human genes for treatment of two of the most common human diseases: induction of gamma-globin gene expression for sickle cell disease and induction of p21 gene expression for cancer. The knowledge obtained from this project will also help understand the molecular mechanisms of PNA binding-induced gene expression and provide direct evidence for treatment of many human diseases including genetic diseases and cancer. The aims are: To determine the molecular mechanisms of PNA-induced gene expression. To investigate parameters that affect PNA delivery efficiency and intracellular PNA binding. To explore the possibility of PNA binding-induced endogenous human gene expression.