Regulation of Polyadenylation: the U1A/U1snRNP Paradigm

多聚腺苷酸化的调控：U1A/U1snRNP 范式

• 批准号: 6825828
• 负责人: SAMUEL I GUNDERSON
• 金额: $ 39.8万
• 依托单位: RUTGERS, THE STATE UNIV OF N.J.
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-01-01 至 2008-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6825828
• 关键词: B lymphocyte; HeLa cells; Papillomavirus; RNA interference; RNA splicing; biotechnology; gene expression; gene induction /repression; genetic techniques; genetically modified animals; keratinocyte; laboratory mouse; microarray technology; nucleic acid sequence; plasmids; polyadenylate; posttranscriptional RNA processing; precursor mRNA; small nuclear RNA; small nuclear ribonucleoproteins; technology /technique development

DESCRIPTION (provided by applicant): Post-transcriptional regulation of the processing of eukaryotic pre-mRNA (splicing, polyadenylation, editing etc.) is an important control point in gene expression in eukaryotes. For nearly all mRNAs, the polyA tail is essential for translation, stability, and nuclear export. Changes in the position of the polyA tail cause changes in the final mRNA sequence leading to additional regulatory possibilities. Regulation of polyA sites, either by alternative polyA site choice or by an "on-off" switch of a single polyA site, was generally considered rare. Remarkably, it is now estimated that 30% of human genes are expressed as mRNAs with different polyA sites, a number likely to be applicable to other vertebrates, as the human genome and EST database are the most complete and best annotated of the vertebrates. Even if only a fraction of this 30% represents regulated polyA sites, we are likely to see a dramatic upward revision in the frequency and importance of this form of regulation. In the first funding period of this grant we: 1) characterized a unique form of polyA tail regulation by the U1A splicing factor, 2) discovered that U1A, via binding non-consensus pre-mRNA sequences, can regulate polyA tail addition to genes other than just to itself and that other proteins are also capable of regulating polyA tail addition in a similar manner, and 3) demonstrated that U1 snRNP can be used as a tool to specifically inhibit expression of specific endogenous genes by inhibiting polyA tail addition. The framework of this proposal builds on work from the first funding period to achieve the following goals: 1) improve/characterize the U1in technique thereby allowing us and others to exploit it to its full potential, 2) investigate a new type of polyA tail regulation involving naturally occurring examples of U1 snRNP inhibiting expression of endogenous genes, and 3) identify/characterize new genes targeted for polyA tail regulation by U1A or by proteins previously shown to have the ability to regulate polyA tail addition. The achievement of these goals will advance fundamental knowledge of mechanisms of regulated gene expression, fully develop a promising new gene inhibitory technique, and elucidate the functional consequences for how components of the splicing machinery nteract with and regulate the polyadenylation machinery.

描述（由申请人提供）：真核前体mRNA加工的转录后调控（剪接、聚腺苷酸化、编辑等）是真核生物基因表达的重要控制点。对于几乎所有的mRNA，polyA尾对于翻译、稳定性和核输出是必不可少的。polyA尾位置的变化导致最终mRNA序列的变化，从而导致额外的调控可能性。通常认为，通过替代polyA位点选择或通过单个polyA位点的“开关”调节polyA位点是罕见的。值得注意的是，现在估计30%的人类基因表达为具有不同polyA位点的mRNA，这一数字可能适用于其他脊椎动物，因为人类基因组和EST数据库是脊椎动物中最完整和注释最好的。即使这30%中只有一小部分代表受调节的polyA位点，我们也可能看到这种形式的调节的频率和重要性急剧上升。在第一个资助期内，我们：1）通过U1 A剪接因子表征了polyA尾调节的独特形式，2）发现U1 A通过结合非共有前mRNA序列可以调节polyA尾添加到基因而不仅仅是其自身，并且其他蛋白质也能够以类似的方式调节polyA尾添加，和3）证明U1 snRNP可用作通过抑制polyA尾添加来特异性抑制特定内源基因表达的工具。本提案的框架以第一个供资期的工作为基础，以实现以下目标：1）改进/表征U1 in技术，从而允许我们和其他人充分利用它的潜力，2）研究一种新型的polyA尾调控，其涉及天然存在的U1 snRNP抑制内源基因表达的实例，和3）鉴定/表征通过U1 A或通过先前显示具有调节polyA尾添加的能力的蛋白质靶向polyA尾调节的新基因。这些目标的实现将推进基因表达调节机制的基础知识，全面开发有前途的新基因抑制技术，并阐明剪接机制的组成部分如何与多聚腺苷酸化机制相互作用并调节多聚腺苷酸化机制的功能后果。