Departments of Biochemistry, Emory University School ofMedicine, Atlanta, Georgia 30322, USA.
J Biol Chem. 2010 Aug 20;285(34):26022-32. doi: 10.1074/jbc.M110.141127. Epub 2010 Jun 16.
Proteins bound to the poly(A) tail of mRNA transcripts, called poly(A)-binding proteins (Pabs), play critical roles in regulating RNA stability, translation, and nuclear export. Like many mRNA-binding proteins that modulate post-transcriptional processing events, assigning specific functions to Pabs is challenging because these processing events are tightly coupled to one another. To investigate the role that a novel class of zinc finger-containing Pabs plays in these coupled processes, we defined the mode of polyadenosine RNA recognition for the conserved Saccharomyces cerevisiae Nab2 protein and assessed in vivo consequences caused by disruption of RNA binding. The polyadenosine RNA recognition domain of Nab2 consists of three tandem Cys-Cys-Cys-His (CCCH) zinc fingers. Cells expressing mutant Nab2 proteins with decreased binding to polyadenosine RNA show growth defects as well as defects in poly(A) tail length but do not accumulate poly(A) RNA in the nucleus. We also demonstrate genetic interactions between mutant nab2 alleles and mutant alleles of the mRNA 3'-end processing machinery. Together, these data provide strong evidence that Nab2 binding to RNA is critical for proper control of poly(A) tail length.
与 mRNA 转录本的多聚（A）尾结合的蛋白质，称为多聚（A）结合蛋白（Pab），在调节 RNA 稳定性、翻译和核输出中发挥着关键作用。像许多调节转录后加工事件的 mRNA 结合蛋白一样，将特定的功能分配给 Pab 是具有挑战性的，因为这些加工事件紧密地耦合在一起。为了研究一类新型含锌指的 Pab 在这些偶联过程中所起的作用，我们定义了保守的酿酒酵母 Nab2 蛋白对多聚腺苷酸 RNA 的识别模式，并评估了 RNA 结合破坏所引起的体内后果。Nab2 的多聚腺苷酸 RNA 识别结构域由三个串联的 Cys-Cys-Cys-His（CCCH）锌指组成。表达与多聚腺苷酸 RNA 结合能力降低的突变 Nab2 蛋白的细胞表现出生长缺陷以及多聚（A）尾长度缺陷，但不会在核内积累多聚（A）RNA。我们还证明了突变 nab2 等位基因与 mRNA 3'端加工机制的突变等位基因之间存在遗传相互作用。这些数据共同提供了强有力的证据，表明 Nab2 与 RNA 的结合对于适当控制多聚（A）尾长度至关重要。
