Profusion of G-quadruplexes on both subunits of metazoan ribosomes

Mammalian and bird ribosomes are nearly twice the mass of prokaryotic ribosomes in part because of their extraordinarily long rRNA tentacles. Human rRNA tentacles are not fully observable in current three-dimensional structures and their conformations remain to be fully resolved. In previous work we identified sequences that favor G-quadruplexes in silico and in vitro in rRNA tentacles of the human large ribosomal subunit. We demonstrated by experiment that these sequences form G-quadruplexes in vitro. Here, using a more recent motif definition, we report additional G-quadruplex sequences on surfaces of both subunits of the human ribosome. The revised sequence definition reveals expansive arrays of potential G-quadruplex sequences on LSU tentacles. In addition, we demonstrate by a variety of experimental methods that fragments of the small subunit rRNA form G-quadruplexes in vitro. Prior to this report rRNA sequences that form G-quadruplexes were confined to the large ribosomal subunit. Our combined results indicate that the surface of the assembled human ribosome contains numerous sequences capable of forming G-quadruplexes on both ribosomal subunits. The data suggest conversion between duplexes and G-quadruplexes in response to association with proteins, ions, or other RNAs. In some systems it seems likely that the integrated population of RNA G-quadruplexes may be dominated by rRNA, which is the most abundant cellular RNA.

哺乳动物与鸟类的核糖体质量近乎为原核生物核糖体的两倍，这在一定程度上源于其拥有异常修长的核糖体RNA（ribosomal RNA, rRNA）触手结构。当前的三维结构解析中尚未完全观测到人类核糖体的rRNA触手，其构象仍有待完整解析。在既往研究中，我们通过计算机模拟（in silico）与体外实验（in vitro），在人类核糖体大亚基的rRNA触手区域筛选出了倾向于形成G-四链体（G-quadruplex）的序列，并通过实验证实这些序列可在体外形成G-四链体。本研究采用更新的基序（motif）定义标准，在人类核糖体两个亚基的表面均发现了额外的G-四链体序列；经修订的序列定义标准显示，大亚基（large subunit, LSU）的触手区域存在广泛分布的潜在G-四链体序列阵列。此外，我们通过多种实验方法证实，核糖体小亚基的rRNA片段可在体外形成G-四链体。在本研究发表前，已被证实可形成G-四链体的rRNA序列仅局限于核糖体大亚基。综合本研究的全部结果可知，组装完成的人类核糖体表面，两个亚基上均存在大量可形成G-四链体的序列。实验数据表明，双链RNA与G-四链体之间可根据与蛋白质、离子或其他RNA的结合状态发生构象转换。在部分细胞体系中，细胞内RNA G-四链体的整体组成可能以rRNA为主——而rRNA正是细胞中丰度最高的RNA。