Expression of snRNA genes

snRNA基因的表达

• 批准号: 6897524
• 负责人: ARNE STENLUND
• 金额: $ 24.58万
• 依托单位: COLD SPRING HARBOR LABORATORY
• 依托单位国家: 美国
• 财政年份: 1987
• 资助国家: 美国
• 起止时间: 1987-07-01 至 2007-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6897524
• 关键词: DNA binding protein; DNA directed RNA polymerase; SDS polyacrylamide gel electrophoresis; chimeric proteins; complementary DNA; enzyme mechanism; gel mobility shift assay; gene expression; genetic mapping; genetic promoter element; genetic transcription; immunoprecipitation; intermolecular interaction; laboratory mouse; laboratory rabbit; monoclonal antibody; nucleoproteins; protein reconstitution; protein structure function; recombinant proteins; small nuclear RNA; tissue /cell culture; transcription factor; western blottings; yeast two hybrid system

DESCRIPTION (provided by applicant): We use the human snRNA genes to study fundamental mechanisms of transcription. They represent an excellent model system for this purpose for a number of reasons. First, snRNA genes contain two types of promoters, one type recognized by RNA polymerase (pol) II and one type recognized by pol III. The pol II and III promoters are very similar in structure and indeed they recruit several common transcription factors. Thus, any difference in the initiation complexes assembled on the two types of promoters is likely to be relevant for the determination of RNA polymerase specificity. Second, these promoters are relatively simple. The core pol II snRNA promoters such as the U1 promoter consist of just one essential element, the proximal sequence element or PSE, which recruits a multisubunit factor called SNAPc. The core pol III snRNA promoters such as the U6 promoter contain a PSE as well as a TATA box, which recruits the TATA box binding protein TBP. Both the pol II and III snRNA promoters contain a distal sequence element or DSE, which activates transcription and contains an octamer sequence that recruits the POU domain protein Oct-1 and a so-called SPH site that recruits the zinc finger protein Staf(ZNF143). Third, we have recently identified a set of well-defined factors required and sufficient for transcription from a pol III snRNA promoter. The identification of this basal pol III transcription machinery allows us to study how it is regulated, and indeed we have shown that CK2 can both activate and repress transcription by phosphorylating different targets within this machinery. The identification of this basal pol lll transcription machinery allows us to study how it is regulated. We propose to 1) continue characterizing the mechanisms of U6 transcription including the architecture of SNAPc, the assembly of the U6 transcription initiation complex, and the impact of Oct-1 on the assembly of this initiation complex; 2) characterize how ZNF143 and Oct-1 cooperate to activate U6 transcription from chromatin templates; and 3) determine how and when CK2 regulates pol III transcription from the human U6 promoter. Together, these experiments will reveal how the U6 transcription complex assembles and specifically recruits pol Ill, and how the assembly and activity of the complex is regulated by activators such as Oct-1 and ZNF143, and by kinases such as CK2, during cell growth and proliferation.

描述（由申请人提供）：我们使用人类SNRNA基因研究转录的基本机制。出于多种原因，它们代表了出色的模型系统。首先，SNRNA基因包含两种类型的启动子，一种由RNA聚合酶（POL）II识别的类型，一种由POL III识别的类型。 Pol II和III启动子在结构上非常相似，实际上它们招募了一些常见的转录因子。因此，在两种类型的启动子上组装的起始复合物的任何差异都可能与测定RNA聚合酶特异性的确定有关。其次，这些启动子相对简单。 U1启动子等核心Pol II snRNA启动子仅由一个必需元素，即近端序列元素或PSE组成，该元件元素或PSE募集了一个称为SNAPC的多基因因子。 U6启动子等核心Pol III SNRNA启动子包含一个PSE和一个TATA盒，该盒募集了TATA盒结合蛋白TBP。 Pol II和III SNRNA启动子都包含远端序列元件或DSE，该元素或DSE激活转录并包含八聚体序列，该序列募集了POU结构域蛋白Oct-1和一个所谓的SPH位点，该位点募集了锌指锌指蛋白STAF（ZNF143）。第三，我们最近确定了一组定义明确的因素，并且足以从POLIII SNRNA启动子进行转录。这种基础POL III转录机制的识别使我们能够研究其调节方式，实际上我们已经表明，CK2可以通过磷酸化该机械中的不同靶标来激活和抑制转录。这种基础波段转录机制的识别使我们能够研究它的调节方式。我们建议1）继续表征U6转录的机制，包括SNAPC的结构，U6转录启动复合物的组装以及OCT-1对该启动复合物组装的影响； 2）表征Znf143和Oct-1如何合作以激活染色质模板的U6转录； 3）确定CK2调节人U6启动子的POL III转录。这些实验将共同揭示U6转录复合物如何组装，并专门募集Pol Ill，以及如何在细胞生长和增殖过程中由Oct-1和ZnF143（例如Oct-1和ZnF143）以及CK2等激酶（例如CK2）调节该复合物的组装和活性。