PLANT INTRON PROCESSING

植物内含子加工

• 批准号: 2179645
• 负责人: Mary A Schuler
• 金额: $ 20.85万
• 依托单位: UNIVERSITY OF ILLINOIS AT URBANA-CHAMPAIGN
• 依托单位国家: 美国
• 财政年份: 1988
• 资助国家: 美国
• 起止时间: 1988-08-26 至 1997-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2179645
• 关键词: RNA biosynthesis; RNA splicing; binding proteins; gel electrophoresis; gene complementation; gene expression; gene mutation; genetic manipulation; genetic models; genetic regulatory element; model design /development; nucleic acid sequence; plant genetics; precursor mRNA; small nuclear RNA; transcription factor

Although it is logical to assume that all eukaryotes share a common mechanism for pre-mRNA splicing, gene transfer experiments indicate that critical differences exist between the events leading to mammalian, yeast and plant intron recognition. Differences which also exist between plant introns have prevented in vivo expression of some plant genes containing introns in transgenic plants. In vivo analysis of the cis-acting factors required for intron recognition in plant nuclei has allowed us to develop a model for intron recognition stating that AU elements spread throughout the length of plant introns roughly define intron boundaries by generating strong AU-transition points and masking internal cryptic sites. Potential splice sites are then selected in a position-dependent manner if they are located upstream (5' splice site) or downstream (3' splice site) from these AU transition points and not if they are embedded within AU-rich intron sequences. This mode of recognition relaxes the need for strong splice site and branchpoint consensus sequences and suggests that plant splicing machineries rely on a variety of novel trans-acting factors. The prominence of AU-rich introns and AU transition points at the intron/exon boundaries of Tetrahymena, Drosophila, C. elegans and S. pombe introns suggests that similar mechanisms for intron recognition and splice site definition may operate in a variety of species. Experiments proposed here are aimed at fully defining cis-acting sequences mediating intron recognition in plant nuclei and isolating trans-acting factors that interact with these sequences. Objectives are: 1) To test the universality of this model for intron recognition on other introns; 2) To define in detail cis-acting elements responsible in vivo for 5' and 3' splice site selection in AU-rich introns; 3) To identify trans-acting factors that interact with the cis-acting sequences, especially AU-binding proteins that appear to differentiate introns from exons in vivo and other proteins that enhance recognition of plant introns in an in vitro HeLa-plant complementation system. These experiments will identify splicing factors and recognition schemes that may mediate excision of AU-rich introns in many species and will determine critical associations that commit pre-mRNAs of this type of a splicing pathway. Ultimately, this information should clarify similarities and differences between the modes of intron recognition operating in mammalian, yeast and plant nuclei and provide for a more thorough mechanistic definition of pre-mRNA splicing.

尽管假设所有真核生物共享一个常见的逻辑 基因转移实验的MRNA剪接机制表明 导致哺乳动物，酵母的事件之间存在关键差异 和植物内含子识别。 植物之间也存在的差异 内含子阻止了某些含有某些植物基因的体内表达 转基因植物的内含子。 在体内分析顺式作用因子 植物核中内含子识别所必需的使我们得以发展 内含子识别的模型表明，AU元素遍布整个 植物内含子的长度大致定义了内含子边界 产生强大的AU转变点并掩盖内部隐秘 站点。 然后选择潜在的剪接位点以位置依赖性 如果它们位于上游（5'剪接站点）或下游（3'）的方式 剪接站点）从这些AU过渡点，而不是嵌入 在富含AU的内含子序列中。 这种识别方式放松了 需要强大的剪接站点和分支点共识序列，以及 表明植物剪接机器依靠各种新颖 跨作用因素。 富含澳元的内含子和au的突出性 过渡点在四膜虫的内含子/外显子边界， 果蝇，秀丽隐杆线虫和S. pombe内含子表明类似 内含子识别和剪接站点定义的机制可能会运行 在多种物种中。 此处提出的实验旨在完全定义顺式作用 介导植物核中内含子识别并分离的序列 与这些序列相互作用的反式作用因子。 目标是： 1）测试该模型的普遍性以在其他方面识别内含子 内含子； 2）详细定义负责体内负责的元素 对于富含Au的内含子的5'和3'剪接位点选择； 3）识别 与顺式作用序列相互作用的反式作用因子， 特别是Au结合蛋白，似乎将内含子与 体内外显子和其他增强植物识别的蛋白质 体外HELA植物互补系统中的内含子。 这些 实验将确定剪接因素和识别方案 可以介导许多物种中富含Au的内含子的切除，并且会 确定关键关联，以使这种类型的A 剪接途径。 最终，这些信息应该澄清 内含子识别模式之间的相似性和差异 在哺乳动物，酵母和植物核中运行，并提供更多 彻底的MRNA剪接的机械定义。