Sir William Dunn School of Pathology, University of Oxford, Oxford OX1 3RE, UK.
Department of Biological and Medical Sciences, Oxford Brookes University, Oxford OX3 0BP, UK.
J Cell Sci. 2022 Oct 1;135(19). doi: 10.1242/jcs.259701. Epub 2022 Oct 4.
The compartmentalised eukaryotic cell demands accurate targeting of proteins to the organelles in which they function, whether membrane-bound (like the nucleus) or non-membrane-bound (like the nucleolus). Nucleolar targeting relies on positively charged localisation signals and has received rejuvenated interest since the widespread recognition of liquid-liquid phase separation (LLPS) as a mechanism contributing to nucleolus formation. Here, we exploit a new genome-wide analysis of protein localisation in the early-branching eukaryote Trypanosoma brucei to analyse general nucleolar protein properties. T. brucei nucleolar proteins have similar properties to those in common model eukaryotes, specifically basic amino acids. Using protein truncations and addition of candidate targeting sequences to proteins, we show both homopolymer runs and distributed basic amino acids give nucleolar partition, further aided by a nuclear localisation signal (NLS). These findings are consistent with phase separation models of nucleolar formation and physical protein properties being a major contributing mechanism for eukaryotic nucleolar targeting, conserved from the last eukaryotic common ancestor. Importantly, cytoplasmic ribosome proteins, unlike mitochondrial ribosome proteins, have more basic residues - pointing to adaptation of physicochemical properties to assist segregation.
真核细胞的区室化要求蛋白质准确靶向其发挥作用的细胞器，无论是膜结合的（如细胞核）还是非膜结合的（如核仁）。核仁靶向依赖于带正电荷的定位信号，并且自从普遍认识到液-液相分离 (LLPS) 是促进核仁形成的机制以来，它重新引起了人们的兴趣。在这里，我们利用对早期分支真核生物布氏锥虫的蛋白质定位的全基因组新分析来分析一般核仁蛋白特性。布氏锥虫核仁蛋白具有与常见模式真核生物相似的特性，特别是碱性氨基酸。通过蛋白质截短和向蛋白质添加候选靶向序列，我们发现无论是同聚氨基酸序列还是分散的碱性氨基酸都可以赋予核仁分配，核定位信号 (NLS) 进一步促进了这一过程。这些发现与核仁形成的相分离模型以及物理蛋白质特性是真核核仁靶向的主要贡献机制一致，这一机制可以追溯到最后一个真核共同祖先。重要的是，细胞质核糖体蛋白与线粒体核糖体蛋白不同，具有更多的碱性残基——这表明物理化学性质的适应性有助于隔离。
