PLANT INTRON PROCESSING

尽管假设所有真核生物都有一个共同的 前mRNA剪接的机制，基因转移实验表明 导致哺乳动物、酵母菌 和植物内含子识别。植物之间也存在的差异 内含子阻止了一些含有以下成分的植物基因的体内表达 转基因植物中的内含子。顺式作用因子的体内分析 在植物核中识别内含子所需的基因使我们能够开发出 一个内含子识别模型表明AU元素分布于 植物内含子的长度通过以下方式大致定义内含子边界 生成强AU-转换点和掩蔽内部隐含 网站。然后以位置相关的方式选择潜在的剪接位点 如果它们位于上游(5‘剪接位点)或下游(3’)，则以 剪接位点)，而不是在它们嵌入的情况下 在富含AU的内含子序列中。这种识别模式放松了 需要强剪接位点和分支点共有序列 建议植物拼接机械依赖于各种新颖的 反式作用因素。富含AU的内含子的重要性和AU 四膜虫内含子/外显子边界的过渡点， 果蝇、线虫和S.pombe内含子表明类似的 用于内含子识别和剪接位点定义的机制可以运行 在不同的物种中。 这里提出的实验旨在充分定义顺式作用 植物核中介导内含子识别的序列及其分离 与这些序列相互作用的反式作用因子。目标是： 1)检验该模型对其他内含子识别的普适性 内含子；2)详细定义体内负责的顺式作用元件 在富含AU的内含子中进行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， 尤其是似乎能区分内含子和内含子的AU结合蛋白 体内的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 实验将识别剪接因子和识别方案， 可能在许多物种中介导富含AU的内含子的切除，并将 确定提交此类型的前mRNA的关键关联 剪接途径。归根结底，这些信息应该澄清 内含子识别模式的异同 在哺乳动物、酵母和植物细胞核中运行，并提供更多 对前信使核糖核酸剪接的机械定义。

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.