Department of Algae Biotechnology & Bioenergy, Faculty of Biology, Bielefeld University, D-33615 Bielefeld, GermanyDepartment of Proteome & Metabolome Research, Faculty of Biology, Bielefeld University, Bielefeld, Germany.
Plant J. 2011 Jan;65(1):119-130. doi: 10.1111/j.1365-313X.2010.04406.x. Epub 2010 Nov 29.
Methylation of protein arginines represents an important post-translational modification mechanism, which has so far primarily been characterized in mammalian cells. In this work, we successfully identified and characterized arginine methylation as a crucial type of post-translational modification in the activity regulation of the cytosolic translation repressor protein NAB1 in the plant model organism Chlamydomonas reinhardtii. NAB1 represses the cytosolic translation of light-harvesting protein encoding mRNAs by sequestration into translationally silent messenger ribonucleoprotein complexes (mRNPs). Protein arginine methylation of NAB1 could be demonstrated by PRMT1 catalyzed methylation of recombinant NAB1 in vitro, and by immunodetection of methylated NAB1 arginines in vivo. Mass spectrometric analyses of NAB1 purified from C. reinhardtii revealed the asymmetric dimethylation of Arg90 and Arg92 within GAR motif I. Inhibition of arginine methylation by either adenosine-2'-3'-dialdehyde (AdOx) or 7,7'-carbonylbis(azanediyl)bis(4-hydroxynaphthalene-2-sulfonic acid) sodium salt hydrate (AMI-1) caused a dark-green phenotype characterized by the increased accumulation of light-harvesting complex proteins, and indicating a reduced translation repressor activity of NAB1. The extent of NAB1 arginine methylation depends on the growth conditions, with phototrophic growth causing a high methylation state and heterotrophic growth resulting in lowered methylation of the protein. In addition, we could show that NAB1 activity regulation by arginine methylation operates independently from cysteine-based redox control, which has previously been shown to control the activity of NAB1.
蛋白质精氨酸的甲基化是一种重要的翻译后修饰机制，迄今为止，这种机制主要在哺乳动物细胞中得到了描述。在这项工作中，我们成功地鉴定并描述了精氨酸甲基化为植物模式生物莱茵衣藻细胞质翻译抑制蛋白 NAB1 活性调节的一种关键的翻译后修饰类型。NAB1 通过将光捕获蛋白编码 mRNA 隔离到翻译沉默的信使核糖核蛋白复合物（mRNP）中来抑制细胞质翻译。通过 PRMT1 体外催化重组 NAB1 的甲基化，可以证明 NAB1 的蛋白精氨酸甲基化，并且可以在体内检测到甲基化的 NAB1 精氨酸。从莱茵衣藻中纯化的 NAB1 的质谱分析表明，GAR 基序 I 内的 Arg90 和 Arg92 发生不对称二甲基化。通过腺嘌呤-2'-3'-二醛（AdOx）或 7,7'-羰基双（氮杂二亚基）双（4-羟基萘-2-磺酸）钠盐水合物（AMI-1）抑制精氨酸甲基化会导致暗绿色表型，其特征是光捕获复合物蛋白的积累增加，表明 NAB1 的翻译抑制活性降低。NAB1 精氨酸甲基化的程度取决于生长条件，光养生长导致高甲基化状态，异养生长导致蛋白质甲基化降低。此外，我们还表明，精氨酸甲基化对 NAB1 活性的调节独立于先前已显示控制 NAB1 活性的半胱氨酸基氧化还原控制。
