Department of Biochemistry, Purdue University, BCHM 305, 175 South University Street, West Lafayette, IN 47907-2063, USA; Purdue University Center for Cancer Research, Purdue University, Hansen Life Sciences Research Building, Room 141, 201 South University Street, West Lafayette, IN 47907-2064, USA.
J Mol Biol. 2013 Oct 23;425(20):3824-38. doi: 10.1016/j.jmb.2013.05.016. Epub 2013 May 28.
Eukaryotic gene expression involves numerous biochemical steps that are dependent on RNA structure and ribonucleoprotein (RNP) complex formation. The DEAD-box class of RNA helicases plays fundamental roles in formation of RNA and RNP structure in every aspect of RNA metabolism. In an effort to explore the diversity of biological roles for DEAD-box proteins, our laboratory recently demonstrated that the DEAD-box protein Dbp2 associates with actively transcribing genes and is required for normal gene expression in Saccharomyces cerevisiae. We now provide evidence that Dbp2 interacts genetically and physically with the mRNA export factor Yra1. In addition, we find that Dbp2 is required for in vivo assembly of mRNA-binding proteins Yra1, Nab2, and Mex67 onto poly(A)+ RNA. Strikingly, we also show that Dbp2 is an efficient RNA helicase in vitro and that Yra1 decreases the efficiency of ATP-dependent duplex unwinding. We provide a model whereby messenger ribonucleoprotein (mRNP) assembly requires Dbp2 unwinding activity and once the mRNP is properly assembled, inhibition by Yra1 prevents further rearrangements. Both Yra1 and Dbp2 are conserved in multicellular eukaryotes, suggesting that this constitutes a broadly conserved mechanism for stepwise assembly of mature mRNPs in the nucleus.
真核生物基因表达涉及许多生化步骤，这些步骤依赖于 RNA 结构和核糖核蛋白 (RNP) 复合物的形成。DEAD -box 类 RNA 解旋酶在 RNA 代谢的各个方面对 RNA 和 RNP 结构的形成起着至关重要的作用。为了探索 DEAD-box 蛋白的生物学作用多样性，我们实验室最近证明 DEAD-box 蛋白 Dbp2 与正在转录的基因结合，并在酿酒酵母中正常基因表达中发挥作用。我们现在提供的证据表明，Dbp2 在遗传和物理上与 mRNA 输出因子 Yra1 相互作用。此外，我们发现 Dbp2 对于体内将 mRNA 结合蛋白 Yra1、Nab2 和 Mex67 组装到 poly(A)+ RNA 上是必需的。引人注目的是，我们还表明 Dbp2 在体外是一种有效的 RNA 解旋酶，并且 Yra1 降低了 ATP 依赖性双链体解旋的效率。我们提供了一个模型，即信使核糖核蛋白 (mRNP) 的组装需要 Dbp2 解旋活性，并且一旦 mRNP 正确组装，Yra1 的抑制作用就会阻止进一步的重排。Yra1 和 Dbp2 在多细胞真核生物中是保守的，这表明这构成了在核中逐步组装成熟 mRNP 的广泛保守机制。
