Structure and function of 5' to 3' exoribonucleases

描述（由申请人提供）：5‘- bbbb3 ’外核糖核酸酶（XRNs）在真核生物中包含一个大家族的保守酶，在RNA代谢和RNA干扰中具有重要功能。在真菌和动物中，XRN1 （175 kD）主要存在于细胞质中，参与去切mrna的降解、无义介导的衰变、microRNA的衰变，是正常发育所必需的。XRN2 （115 kD）主要是细胞核，其在酿酒酵母中的同源物，通常被称为Rat1，在RNA从细胞核转运到细胞质和RNA聚合酶III转录一些基因中起作用。最近的研究已经确定XRN2/Rat1是RNA聚合酶II （Pol II）转录终止的“鱼雷”模型所必需的外核糖核酸酶。在酿酒酵母中，Rat1与另一种蛋白Rai1 （Rat1相互作用蛋白1）共纯化，这似乎增强了Rat1的体外核糖核酸酶活性。Rai1的弱同源序列DOM3Z存在于哺乳动物中，尽管它可能与XRN2没有关联。XRNs在RNA代谢、RNA干扰和Pol II终止中的重要功能引起了人们的极大兴趣。然而，这些酶的功能的分子机制目前仍然知之甚少。作为填补这些知识空白的第一步，我们最近确定了Rat1-Rai1复合物以及Rai1和DOM3Z单独的晶体结构，并根据结构信息进行了初步的生化和功能研究。这些结果为进一步研究这些蛋白质的结构、生化和生物学奠定了基础。最有趣的是，我们的初步研究表明，Rai1/DOM3Z含有一个单独的活性位点，具有RNA 5'焦磷酸水解酶活性，我们假设Rai1/DOM3Z可能在RNA代谢或质量控制中具有重要功能。我们还将在我们提出的研究项目中验证这一假设，这将有助于确定这种新型真核酶的生理功能。

DESCRIPTION (provided by applicant): The 5'->3' exoribonucleases (XRNs) comprise a large family of conserved enzymes in eukaryotes with important functions in RNA metabolism and RNA interference. In fungi and animals, XRN1 (175 kD) is primarily cytosolic and is involved in degradation of decapped mRNAs, nonsense mediated decay, microRNA decay and is essential for proper development. XRN2 (115 kD) is primarily nuclear, and its ortholog in S. cerevisiae, more commonly known as Rat1, has roles in RNA trafficking from the nucleus to the cytoplasm and transcription of some genes by RNA polymerase III. More recent studies have identified XRN2/Rat1 as the exoribonuclease that is essential for the 'torpedo' model of transcription termination by RNA polymerase II (Pol II). In S. cerevisiae, Rat1 co-purifies with another protein, Rai1 (Rat1 interacting protein 1), which appears to enhance the exoribonuclease activity of Rat1 in vitro. A weak sequence homolog of Rai1, known as DOM3Z, exists in mammals, although it probably does not associate with XRN2. The important functions of the XRNs in RNA metabolism, RNA interference and Pol II termination have generated significant interest in them. However, the molecular mechanism for the functions of these enzymes is currently still poorly understood. As a first step to fill these gaps in our knowledge, we recently determined the crystal structures of the Rat1-Rai1 complex as well as of Rai1 and DOM3Z alone, and carried out preliminary biochemical and functional studies based on the structural information. These results set the stage for further structural, biochemical and biological studies on these proteins. Most interestingly, our initial studies show that Rai1/DOM3Z contains a separate active site, with RNA 5' pyrophosphohydrolase activity, and we hypothesize that Rai1/DOM3Z may have important functions in RNA metabolism or quality control. We will also test this hypothesis in our proposed research project, which will help to define the physiological functions of this novel eukaryotic enzyme.