PLANT INTRON PROCESSING

虽然逻辑上可以假设所有的真核生物都有一个共同的 前体mRNA剪接的机制，基因转移实验表明， 在导致哺乳动物、酵母菌、 和植物内含子识别。 植物之间也存在差异 内含子已经阻止了一些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 在植物细胞核中识别内含子所需的条件，使我们能够开发 内含子识别的模型表明Au元素遍布 植物内含子的长度大致限定了内含子边界， 产生强AU-转移点和掩蔽内部隐藏 网站. 然后以位置依赖性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 ′剪接位点）， 剪接位点），而不是如果它们被嵌入 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， 表明植物拼接机器依赖于各种新颖的 反式作用因子 富含AU的内含子和Au的突出 在四膜虫的内含子/外显子边界处的转换点， 果蝇C. elegans和S.粟酒属内含子表明， 内含子识别和剪接位点定义的机制可能会起作用 在不同的物种中。 这里提出的实验旨在充分定义顺式作用 介导植物细胞核中内含子识别的序列和分离 与这些序列相互作用的反式作用因子。 目标是： 1)为了检验该模型在其他基因组上内含子识别的通用性， 内含子; 2）详细定义在体内起作用的顺式作用元件 用于富含AU的内含子中的5'和3'剪接位点选择; 3）鉴定 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，它们似乎将内含子与 体内外显子和其他增强植物识别的蛋白质 在体外HeLa-植物互补系统中的内含子。 这些 实验将确定剪接因子和识别方案， 可能介导许多物种中富含AU的内含子的切除， 确定提交这种类型的前mRNA的关键关联， 剪接途径 最终，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， 前体mRNA剪接的彻底机制定义。

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.