GENETIC ANALYSIS OF TRNA SPLICING IN THE YEAST NUCLEUS

从真核前体-tRNAs剪接插入序列 三种酶的作用，一种是异三聚体核酸内切酶，一种是 多功能但单体连接酶，以及NAD连接的磷酸酶。 核酸内切酶和连接酶形成一个物理复合体，定位于 核孔附近的核周，表明 剪接和核出口的功能耦合的潜力。二 此前在酵母中已鉴定出SEN1和SEN2基因 酵母菌是核酸内切酶活性所必需的。 核定位的核酸内切酶阳性效应子的SEN1编码 似乎不是酶的催化亚基的活性。SEN2 该酶的42kD催化亚基的编码。我们已经隔离了 影响SEN1功能的六个新基因的突变。两个基因突变 其中，SEN3和SEN4基因影响SEN1的表达 翻译后水平。一个温度敏感突变，sen3-L， 也会导致体内tRNA剪接中间产物的过度积累。 此外，已经确定了四个基因，它们抑制了 温度敏感型SEL-L突变存在于多个拷贝和 可能编码与SEN1蛋白相互作用的产物。通过继续 通过我们对SEN1和SEN2功能的分析以及通过启动分析 SEN3、SEN4和高拷贝抑制基因，我们希望提供 关于结构、组织、功能和 剪接复合体的细胞内位置。也有可能是 我们可能会学到一些关于剪接酶引入到 细胞核和从细胞核输出剪接的tRNA产物。

Intervening sequences are spliced from eukaryotic precursor-tRNAs through the action of three enzymes, a heterotrimeric endonuclease, a multi-functional but monomeric ligase, and an NAD-linked phosphatase. The endonuclease and ligase form a physical complex that is localized in the nuclear periphery in the vicinity of nuclear pores, suggesting the potential for functional coupling of splicing and nuclear export. Two genes, SEN1 and SEN2, were identified previously in the yeast Saccharomyces cerevisiae that are required for endonuclease activity. SEN1 codes for a nuclear-localized, positive effector of endonuclease activity that appears not to be a catalytic subunit of the enzyme. SEN2 codes for the 42 Kd catalytic subunit of the enzyme. We have isolated mutations in six new genes that affect SEN1 function. Mutations in two of the genes, SEN3 and SEN4, affect the expression of SEN1 at a post-translational level. One temperature-sensitive mutation, sen3-l, also causes excess accumulation of tRNA splicing intermediates in vivo. In addition, four genes have been identified that suppress the temperature-sensitive senl-l mutation when present in multiple copies and may encode products that interact with the SEN1 protein. By continuing with our analysis of SEN1 and SEN2 function and by initiating analyses of SEN3, SEN4, and the high-copy suppressor genes, we hope to provide critical information on the structure, organization, function, and intracellular location of the splicing complex. It is also possible that we may learn something about the import of splicing enzymes into the nucleus and the export of spliced tRNA products from the nucleus.