Eukaryote-specific rRNA expansion segments function in ribosome biogenesis

Abstract

The secondary structure of ribosomal RNA (rRNA) is largely conserved across all kingdoms of life. However, eukaryotes have evolved extra blocks of rRNA sequences, relative to those of prokaryotes, called expansion segments (ES). A thorough characterization of the potential roles of ES remains to be done, possibly because of limitations in the availability of robust systems to study rRNA mutants. We sought to systematically investigate the potential functions, if any, of the ES in 25S rRNA of Saccharomyces cerevisiae by deletion mutagenesis. We deleted 14 of the 16 different eukaryote-specific ES in yeast 25S rRNA individually and assayed their phenotypes. Our results show that all but two of the ES tested are necessary for optimal growth and are required for production of 25S rRNA, suggesting that ES play roles in ribosome biogenesis. Further, we classified expansion segments into groups that participate in early nucleolar, middle, and late nucleoplasmic steps of ribosome biogenesis, by assaying their pre-rRNA processing phenotypes. This study is the first of its kind to systematically identify the functions of eukaryote-specific expansion segments by showing that they play roles in specific steps of ribosome biogenesis. The catalog of phenotypes we identified, combined with previous investigations of the roles ribosomal proteins in large subunit biogenesis, leads us to infer that assembling ribosomes are composed of distinct RNA and protein structural neighborhood clusters that participate in specific steps of ribosome biogenesis.

Size-variable zone in V3 region of 16S rRNA

“…H17, together with H16 and H18, are considered parts of regions characterized as Expansion Segments, where functional insertions have been registered. 32,33 Further study on rRNA structure may help better explain this event.…”

Expression of distinct maternal and somatic 5.8S, 18S, and 28S rRNA types during zebrafish development

“…ESs exhibit a significant degree of variability between species in length and sequence, plus their size and number of helical branches seem to progressively increase in higher eukaryotes (Michot and Bachellerie 1987). The role of the 28S ESs in ribosomal function is still not clear, but it has recently been shown that they are important in specific steps of the ribosome biogenesis in yeast (Ramesh and Woolford 2016). The many differences we observe between the ESs of maternal-and somatic-type 28S will undoubtedly have an effect on assembly and functioning of the ribosomal LSU.…”

Structure and assembly model for the Trypanosoma cruzi 60S ribosomal subunit

“…Thus, the arrangement of the srRNAs as well as their anchoring proteins help define the likely order of assembly of the srRNAs, such that stable association of srRNA2 and srRNA3 occurs before that of srRNA1 and srRNA4. Indeed, deletion of the counterpart of srRNA3 in yeast (ES39L) leads to an early processing defect (19). Strikingly, although srRNA1 is the first small rRNA transcribed, it is stably assembled after srRNA2 and srRNA3, suggesting that its RNA processing occurs during or after assembly of the ribosome instead of the 5′ to 3′ direction following transcription.…”

“…This helix contacts extensions of eL8 and interacts with the 3′-end of the 5.8S rRNA. In yeast, deleting ES31L or the N-terminal extension of eL8 blocks ribosome assembly at later stages, suggesting that ES31L may be involved in subsequent stabilization steps of the previously established scaffold (23). Also associated with the later stages is the final positioning of the 5S rRNA.…”