Regulation of mRNA Stability in Plastids - a Genetic and Biochemical Approach

质体中 mRNA 稳定性的调控——一种遗传和生化方法

• 批准号: 9404841
• 负责人: Wilhelm Gruissem
• 金额: $ 29.8万
• 依托单位: University of California-Berkeley
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 1994
• 资助国家: 美国
• 起止时间: 1994-07-15 至 1997-06-30
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=9404841&HistoricalAwards=false
• 关键词: Regulation; mRNA; Stability; Plastids; Genetic

9404841 Gruissem Plastids in higher plants respond to the developmental program and environmental signals by adjusting the transcription rate of genes and the processing and stability of their mRNAs. The change in plastid mRNA stability is an important regulatory mechanism that is of functional consequence for translational efficiency. Most plastid mRNAs have an inverted repeat (IR) sequence in their 3' untranslated region (UTR) that is required for correct mRNA 3' end processing and the stability of the mature mRNA. Mutagenesis or deletion of the 3' IR sequence causes incorrect processing and/or rapid decay of the mRNA. Five nuclear-encoded proteins have been identified that interact with the 3' UTR of several plastid mRNAs in a structural and/or sequence-dependent manner. This project will take a combined biochemical and genetic approach to investigate the function of each protein and their possible interactions. Al proteins will be tested individually or in combinations for their role in mRNA 3' end processing and stability using reconstituted or partially fractionated in vitro systems. These studies will be complemented by parallel antisense-RNA strategies using Arabidopsis to confirm the in vivo function of the proteins. Previous experiments have already shown that the 28RNP is critical for chloroplast development in leaves. Similar antisene-RNA strategies will be used to determine the developmental role of the 24RNP which differs in its expression pattern from the 28RNP. The 100RNP will be tested for its potential interaction with the 24RNP and 28RNP to form a mRNA processing complex. Other experiments will investigate the function of the 29RNP and 55RNP and the interaction of these proteins with the 3'UTR of the petD mRNA. Together the biochemical and genetic dissection of the RNA-binding proteins will provide novel information on the nuclear control of organelle gene expression. Because of the importance of photosynthesis for plant productivity, the basic research proposed in this application is likely to have relevance for future plant and agricultural technologies. %%% Development of chloroplasts and photosynthetic competence are critical for plant growth and function. It has been shown that plastids respond to the plant developmental program and environmental signals by adjusting the expression of genes encoded in the plastid genome. One important control of plastid gene expression during development occurs at the level of mRNA processing and stability. Changes in mRNA stability can directly affect the translation of proteins that have essential functions in photosynthesis and other chloroplast processes. Processing and stability of chloroplast mRNAs are mediated by nuclear-encoded proteins that interact with specific sequences or structures in the RNA molecule. In previous studies, five proteins have been identified that participate in the processing and stability of a specific class of chloroplast mRNAs. The current project will take a combined biochemical and genetic approach to investigate the function of each protein and their possible interactions. Together, the biochemical and genetic dissection of the mechanisms by which chloroplast mRNA stability is accomplished will provide novel information on the nuclear control of organelle gene expression. Because of the importance of photosynthesis for plant productivity, the basic research proposed in this project is likely to have relevance for future plant and agricultural technologies. ***

9404841高等植物中的Gruissem质体通过调节基因的转录速率及其mRNA的加工和稳定性来响应发育程序和环境信号。 质体mRNA稳定性的变化是重要的调控机制，其对翻译效率具有功能性后果。 大多数质体mRNA在其3'非翻译区（UTR）中具有反向重复（IR）序列，其是正确mRNA 3'末端加工和成熟mRNA的稳定性所需的。 3' IR序列的突变或缺失导致mRNA的不正确加工和/或快速衰减。 已经鉴定了五种核编码蛋白，其以结构和/或序列依赖性方式与几种质体mRNA的3' UTR相互作用。 该项目将采用生物化学和遗传学相结合的方法来研究每种蛋白质的功能及其可能的相互作用。 将使用重构或部分分级分离的体外系统单独或组合测试Al蛋白在mRNA 3'末端加工和稳定性中的作用。 这些研究将通过使用拟南芥的平行反义RNA策略来补充，以确认蛋白质的体内功能。 先前的实验已经表明，28 RNP对叶片中的叶绿体发育至关重要。 类似的反义RNA策略将用于确定24 RNP的发育作用，其表达模式不同于28 RNP。 将测试100 RNP与24 RNP和28 RNP形成mRNA加工复合物的潜在相互作用。 其他实验将研究29 RNP和55 RNP的功能以及这些蛋白质与petD mRNA的3 'UTR的相互作用。 RNA结合蛋白的生物化学和遗传学解剖将为细胞器基因表达的核控制提供新的信息。 由于光合作用对植物生产力的重要性，本申请中提出的基础研究可能与未来的植物和农业技术有关。 叶绿体的发育和光合能力对植物的生长和功能至关重要。 研究表明，质体通过调节质体基因组中编码基因的表达来响应植物发育程序和环境信号。 发育过程中质体基因表达的一个重要控制发生在mRNA加工和稳定性水平。 mRNA稳定性的变化可以直接影响在光合作用和其他叶绿体过程中具有重要功能的蛋白质的翻译。 叶绿体mRNA的加工和稳定性是由核编码的蛋白质介导的，这些蛋白质与RNA分子中的特定序列或结构相互作用。 在以前的研究中，已经确定了五种蛋白质参与一类特定叶绿体mRNA的加工和稳定性。 目前的项目将采取生物化学和遗传学相结合的方法来研究每种蛋白质的功能及其可能的相互作用。 总之，生化和遗传解剖的机制，叶绿体mRNA的稳定性完成将提供新的信息细胞器基因表达的核控制。 由于光合作用对植物生产力的重要性，本项目提出的基础研究可能与未来的植物和农业技术有关。 ***