Departamento de Genética, Universidad de Sevilla, E-41012 Seville, Spain, Institut Pasteur, Génétique des Interactions Macromoléculaires, CNRS UMR-3525, Paris, France and Instituto de Bioquímica Vegetal y Fotosíntesis, Universidad de Sevilla-CSIC, Seville, Spain.
Nucleic Acids Res. 2013 Nov;41(20):9461-70. doi: 10.1093/nar/gkt726. Epub 2013 Aug 13.
Ribosome biogenesis requires >300 assembly factors in Saccharomyces cerevisiae. Ribosome assembly factors Imp3, Mrt4, Rlp7 and Rlp24 have sequence similarity to ribosomal proteins S9, P0, L7 and L24, suggesting that these pre-ribosomal factors could be placeholders that prevent premature assembly of the corresponding ribosomal proteins to nascent ribosomes. However, we found L7 to be a highly specific component of Rlp7-associated complexes, revealing that the two proteins can bind simultaneously to pre-ribosomal particles. Cross-linking and cDNA analysis experiments showed that Rlp7 binds to the ITS2 region of 27S pre-rRNAs, at two sites, in helix III and in a region adjacent to the pre-rRNA processing sites C1 and E. However, L7 binds to mature 25S and 5S rRNAs and cross-linked predominantly to helix ES7(L)b within 25S rRNA. Thus, despite their predicted structural similarity, our data show that Rlp7 and L7 clearly bind at different positions on the same pre-60S particles. Our results also suggest that Rlp7 facilitates the formation of the hairpin structure of ITS2 during 60S ribosomal subunit maturation.
核糖体生物发生需要 >300 种装配因子在酿酒酵母中。核糖体装配因子 Imp3、Mrt4、Rlp7 和 Rlp24 与核糖体蛋白 S9、P0、L7 和 L24 具有序列相似性，表明这些前核糖体因子可能是占位符，可防止相应的核糖体蛋白过早地装配到新生核糖体上。然而，我们发现 L7 是 Rlp7 相关复合物的高度特异性成分，这表明这两种蛋白可以同时结合到前核糖体颗粒上。交联和 cDNA 分析实验表明，Rlp7 结合到 27S 前 rRNA 的 ITS2 区域，在两个位点，在螺旋 III 和在靠近 pre-rRNA 加工位点 C1 和 E 的区域。然而，L7 结合到成熟的 25S 和 5S rRNA，并主要交联到 25S rRNA 内的 ES7(L)b 螺旋。因此，尽管它们具有预测的结构相似性，但我们的数据表明，Rlp7 和 L7 在前 60S 颗粒上的不同位置明显结合。我们的结果还表明，Rlp7 有助于在 60S 核糖体亚基成熟过程中形成 ITS2 的发夹结构。
