a Institute of Biochemistry and Biotechnology , Martin Luther University Halle-Wittenberg , Halle , Germany.
RNA Biol. 2019 Jul;16(7):960-971. doi: 10.1080/15476286.2019.1602438. Epub 2019 Apr 15.
The RNA-binding protein AUF1 regulates post-transcriptional gene expression by affecting the steady state and translation levels of numerous target RNAs. Remodeling of RNA structures by the largest isoform AUF1 p45 was recently demonstrated in the context of replicating RNA viruses, and involves two RNA remodeling activities, i.e. an RNA chaperone and an RNA annealing activity. AUF1 contains two non-identical RNA recognition motifs (RRM) and one RGG/RG motif located in the C-terminus. In order to determine the functional significance of each motif to AUF1's RNA-binding and remodeling activities we performed a comprehensive mutagenesis study and characterized the wildtype AUF1, and several variants thereof. We demonstrate that each motif contributes to efficient RNA binding and remodeling by AUF1 indicating a tight cooperation of the RRMs and the RGG/RG motif. Interestingly, the data identify two distinct roles for the arginine residues of the RGG/RG motif for each RNA remodeling activity. First, arginine-mediated stacking interactions promote AUF1's helix-destabilizing RNA chaperone activity. Second, the electropositive character of the arginine residues is the major driving force for the RNA annealing activity. Thus, we provide the first evidence that arginine residues of an RGG/RG motif contribute to the mechanism of RNA annealing and RNA chaperoning.
AUF1 是一种 RNA 结合蛋白，通过影响众多靶 RNA 的稳态和翻译水平来调节转录后基因表达。最近在复制 RNA 病毒的背景下证明了最大的异构体 AUF1 p45 对 RNA 结构的重塑，这涉及两种 RNA 重塑活性，即 RNA 伴侣和 RNA 退火活性。AUF1 包含两个非同源的 RNA 识别基序（RRM）和一个位于 C 末端的 RGG/RG 基序。为了确定每个基序对 AUF1 的 RNA 结合和重塑活性的功能意义，我们进行了全面的突变研究，并对野生型 AUF1 及其几种变体进行了表征。我们证明，每个基序都有助于 AUF1 有效地结合和重塑 RNA，表明 RRMs 和 RGG/RG 基序之间的紧密合作。有趣的是，这些数据确定了 RGG/RG 基序中的两个精氨酸残基在每种 RNA 重塑活性中的两个不同作用。首先，精氨酸介导的堆积相互作用促进了 AUF1 的螺旋解稳定 RNA 伴侣活性。其次，精氨酸残基的正电性是 RNA 退火活性的主要驱动力。因此，我们首次提供了证据，证明 RGG/RG 基序中的精氨酸残基有助于 RNA 退火和 RNA 伴侣的机制。
