Fernández-Pevida Antonio, Martín-Villanueva Sara, Murat Guillaume, Lacombe Thierry, Kressler Dieter, de la Cruz Jesús
Instituto de Biomedicina de Sevilla (IBiS), Hospital Universitario Virgen del Rocío/CSIC/Universidad de Sevilla, Avda. Manuel Siurot, s/n; E-41013 Seville, Spain Departamento de Genética, Universidad de Sevilla, Seville, Spain.
Unit of Biochemistry, Department of Biology, University of Fribourg, Chemin du Musée 10, CH-1700 Fribourg, Switzerland.
Nucleic Acids Res. 2016 Sep 19;44(16):7777-91. doi: 10.1093/nar/gkw641. Epub 2016 Jul 15.
The archaea-/eukaryote-specific 40S-ribosomal-subunit protein S31 is expressed as an ubiquitin fusion protein in eukaryotes and consists of a conserved body and a eukaryote-specific N-terminal extension. In yeast, S31 is a practically essential protein, which is required for cytoplasmic 20S pre-rRNA maturation. Here, we have studied the role of the N-terminal extension of the yeast S31 protein. We show that deletion of this extension partially impairs cell growth and 40S subunit biogenesis and confers hypersensitivity to aminoglycoside antibiotics. Moreover, the extension harbours a nuclear localization signal that promotes active nuclear import of S31, which associates with pre-ribosomal particles in the nucleus. In the absence of the extension, truncated S31 inefficiently assembles into pre-40S particles and two subpopulations of mature small subunits, one lacking and another one containing truncated S31, can be identified. Plasmid-driven overexpression of truncated S31 partially suppresses the growth and ribosome biogenesis defects but, conversely, slightly enhances the hypersensitivity to aminoglycosides. Altogether, these results indicate that the N-terminal extension facilitates the assembly of S31 into pre-40S particles and contributes to the optimal translational activity of mature 40S subunits but has only a minor role in cytoplasmic cleavage of 20S pre-rRNA at site D.
古细菌/真核生物特有的40S核糖体亚基蛋白S31在真核生物中作为泛素融合蛋白表达，由一个保守的主体和一个真核生物特有的N端延伸部分组成。在酵母中，S31是一种几乎必不可少的蛋白质，是细胞质20S前体rRNA成熟所必需的。在这里，我们研究了酵母S31蛋白N端延伸部分的作用。我们发现，删除该延伸部分会部分损害细胞生长和40S亚基生物合成，并导致对氨基糖苷类抗生素超敏。此外，该延伸部分含有一个核定位信号，可促进S31的主动核输入，S31与细胞核中的前核糖体颗粒相关联。在没有该延伸部分的情况下，截短的S31不能有效地组装成前40S颗粒，并且可以鉴定出成熟小亚基的两个亚群，一个缺乏截短的S31，另一个含有截短的S31。质粒驱动的截短S31过表达部分抑制了生长和核糖体生物合成缺陷，但相反，略微增强了对氨基糖苷类的超敏性。总之，这些结果表明，N端延伸部分促进S31组装成前40S颗粒，并有助于成熟40S亚基的最佳翻译活性，但在20S前体rRNA在D位点的细胞质切割中仅起次要作用。
