Upf1p, Nmd2p, and Upf3p are interacting components of the yeast nonsense-mediated mRNA decay pathway
Upf1p, Nmd2p, and Upf3p are interacting components of the yeast nonsense-mediated mRNA decay pathway
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DOI：
10.1128/mcb.17.3.1580
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发表时间：
1997-03-01
影响因子：
5.3
通讯作者：
Jacobson, A
中科院分区：
文献类型：
--
作者：
He, F;Brown, AH;Jacobson, A
酿酒酵母中无义mRNA的快速周转依赖于Upf 1 p、Nmd 2 p和Upf 3 p，它们分别是UPF 1、NMD 2/UPF 2和UPF 3基因的产物。我们先前表明Upf 1 p和Nmd 2 p相互作用，并且这种相互作用是无义介导的mRNA衰变所必需的（F，He和A，Jacobson，Genes Dev，9：437-454，1995; F，He，A，H，Brown和A，Jacobson，RNA 2：153-170，1996），在这项研究中，我们使用酵母双杂交系统来确定该衰变途径的基本组分中的其他蛋白质-蛋白质相互作用，鉴定了Nmd 2 p-Upf 3 p和Upf 1 p-Upf 3 p相互作用，通过缺失分析确定了参与相互作用的相应结构域，发现Upf 1 p和Upf 3 p相互作用的结构域对应于与Nmd 2 p相互作用的两种蛋白质上的结构域，表明Nmd 2 p桥接Upf 1 p和Upf 3 p，该结论通过实验得到加强，表明：（i）NMD 2的缺失完全消除了Upf 1 p和Upf 3 p之间的相互作用，和（ii）保留Upf 1 p-和Upf 3 p-相互作用结构域的全长Nmd 2 p或Nmd 2 p片段的过表达促进了Upf 1 p-Nmd 2 p-Upf 3 p复合物形成的10- 200倍增强。观察到具有UPF 1、NMD 2和UPF 3的单个或多个缺失的细胞以相同程度抑制无义介导的mRNA衰减;并且对显性阴性NMD 2等位基因的可能靶点的分析表明Upf 1 p、Nmd 2 p、Upf 3 p和至少一种其它因子是功能依赖性的，酵母无义介导的mRNA衰变途径的相互作用组分。
Rapid turnover of nonsense-containing mRNAs in Saccharomyces cerevisiae is dependent on Upf1p, Nmd2p, and Upf3p, the products of the UPF1, NMD2/UPF2, and UPF3 genes, respectively, We showed previously that Upf1p and Nmd2p interact and that this interaction is required for nonsense-mediated mRNA decay (F, He and A, Jacobson, Genes Dev, 9:437-454, 1995; F, He, A, H, Brown, and A, Jacobson, RNA 2:153-170, 1996), In this study we have used the yeast two-hybrid system to define other protein-protein interactions among the essential components of this decay pathway, Nmd2p-Upf3p and Upf1p-Upf3p interactions were identified, and the respective domains involved in these interactions were delineated by deletion analysis, The domains of Upf1p and Upf3p putatively involved in their mutual interaction were found to correspond to the domains on the two proteins which interact with Nmd2p, suggesting that Nmd2p bridges Upf1p and Upf3p, This conclusion was reinforced by experiments showing that: (i) deletion of NMD2 completely abolishes interactions between Upf1p and Upf3p and (ii) overexpression of full-length Nmd2p or Nmd2p fragments that retain Upf1p- and Upf3p-interacting domains promotes 10- to 200-fold enhancement of Upf1p-Nmd2p-Upf3p complex formation, These results; the observation that cells harboring either single or multiple deletions of UPF1, NMD2, and UPF3 inhibit nonsense-mediated mRNA decay to the same extent; and an analysis of the possible targets of a dominant-negative NMD2 allele indicate that Upf1p, Nmd2p, Upf3p, and at least one other factor are functionally dependent, interacting components of the yeast nonsense-mediated mRNA decay pathway.