REGULATION OF MESSENGER RNA SPLICING

信使 RNA 剪接的调控

• 批准号: 6410161
• 负责人: Adrian R Krainer
• 金额: $ 22.84万
• 依托单位: COLD SPRING HARBOR LABORATORY
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-01-01 至 2001-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6410161
• 关键词: HeLa cells; RNA binding protein; RNA splicing; X ray crystallography; cell transformation; chimeric proteins; chromatography; genetic regulation; heterogeneous nuclear ribonucleoprotein; immunoprecipitation; laboratory mouse; laboratory rabbit; messenger RNA; molecular cloning; molecular oncology; monoclonal antibody; oncogenes; posttranslational modifications; protein purification; protein structure function; transcription factor; virus genetics; western blottings

The control of gene expression at the post-transcriptional level is a fundamental problem in biology, with relevance to cancer. The mechanisms for the regulation of alternative splicing of cellular and viral genes will be investigated, focusing on global mechanisms that affect the expression of large sets of pre -mRNAs in different tissues, developmental stages, and/or in response to external signals. Three families of human alternative splicing factors will be investigated: (I) the SF7 factors, exemplified by the recently identified SF7A protein; (ii) the hnRNP A/B proteins, of which the best characterized is hnRNP A1; and (iii) the SR proteins, whose prototype is SF2/ASF. The SR proteins function also as constitutive splicing factors, but this project will focus on their ability to modulate alternative splicing in vivo and in vitro in a concentration- dependent manner. This activity is antagonized by hnRNP A/B proteins to modulate alternative 5' splice site selection, and by SF7 proteins to determine alternative 3' splice site selection. The molecular mechanisms by which individual members of these families of RNA-binding proteins modulate alternative splice site selection, and how they achieve substrate specificity, will continue to be studies using biochemical, molecular, and reverse genetic approaches. In addition, the hypothesis that specific pairwise combinations of these antagonistic factors are used to regulate alternative splicing of specific sets of pre-mRNAs will be tested, and experiments are proposed to identify natural, specific pre-mRNA targets for regulations by each of these proteins in vivo. These studies are directly relevant to the overall goals of the program project. As global regulators of alternative pre-mRNA splicing, the SR, hnRNP A/B, and SF7 proteins are excellent candidates to account for the observed aberrant patterns of mRNA expression of numerous genes in transformed cells. Among potential targets of these regulators are several critical genes involved in the establishment or maintenance of the transformed phenotype, or in progression of malignancy. Regulation of alternative splicing is responsible for generating oncongenic and non-oncognic forms of many cellular and viral oncogenes. Therefore, a abetter understanding of the basic mechanisms of alternative splicing regulation, and of the specificity of this process, may lead, in the long term, to the identification of drugs that specifically affect the synthesis of particular protein isoforms that play critical roles in tumorigenesis.

转录后水平的基因表达控制是 生物学中与癌症相关的基本问题。 这 细胞选择性剪接的调节机制 将研究病毒基因，重点关注全球机制 影响不同细胞中大量前体 mRNA 的表达 组织、发育阶段和/或对外部信号的反应。 人类选择性剪接因子的三个家族将是 调查：(I) SF7 因素，例如最近 鉴定出SF7A蛋白； (ii) hnRNP A/B 蛋白，其中 最具特征的是 hnRNP A1； (iii) SR 蛋白，其 原型是SF2/ASF。 SR 蛋白也可作为组成型蛋白 剪接因素，但该项目将重点关注它们的能力 在体内和体外以一定浓度调节选择性剪接- 依赖方式。 该活性被 hnRNP A/B 拮抗 调节选择性 5' 剪接位点选择的蛋白质，以及 SF7 蛋白质以确定替代 3' 剪接位点选择。这 这些个体成员的分子机制 RNA 结合蛋白家族调节可变剪接位点 选择以及它们如何实现底物特异性，将继续 使用生物化学、分子和反向遗传学进行研究 接近。 此外，假设特定的成对 这些拮抗因素的组合被用来调节 将测试特定前体 mRNA 组的选择性剪接，并且 建议进行实验来鉴定天然的、特异性的前 mRNA 这些蛋白质在体内的调节目标。这些 研究与该计划的总体目标直接相关 项目。 作为选择性前 mRNA 剪接的全球调节者， SR、hnRNP A/B 和 SF7 蛋白是 解释观察到的 mRNA 表达的异常模式 转化细胞中的大量基因。潜在目标中 这些调节因子是参与调节的几个关键基因 转化表型的建立或维持，或 恶性肿瘤的进展。 选择性剪接的调节是 负责产生致癌和非致癌形式 许多细胞和病毒癌基因。 因此，一个更好的 了解选择性剪接的基本机制 监管以及这一过程的特殊性可能会导致 从长远来看，要确定特别影响的药物 特定蛋白质亚型的合成，在 肿瘤发生。