Poster abstracts
Poster number 60 submitted by Stefanie Hedayati
The role of IDRs in the assembly pathway of pre-ribosomes
Stefanie A. Hedayati (Department of Biological Sciences, Carnegie Mellon University), Collin Bachert (Department of Biological Sciences, Carnegie Mellon University), John L. Woolford (Department of Biological Sciences, Carnegie Mellon University)
Abstract:
Eukaryotic ribosome biogenesis occurs in a step-wise fashion in the nucleolus, nucleoplasm and cytoplasm. About 200 assembly factors (AFs) are required for the maturation of both the large and small subunit. Cryo-EM structures of co-transcriptional to cytoplasmic pre-ribosome intermediates have provided structural information about the composition and conformation of assembly factors, ribosomal proteins and compacted rRNA in these complexes. However, many AFs contain intrinsically disordered regions that are not visualized by cryo-EM. Interestingly, a major fraction of IDR-containing AFs is released right when pre-ribosomes transit from the nucleolus to the nucleoplasm. We hypothesize that these IDRs contribute to form the nucleolar condensate, a phase-separated non-membrane bound compartment, via trans interactions between pre-ribosomal particles, and that the release of these IDR-containing AFs triggers the nucleolar exit of pre-ribosomes. Consistent with this idea, mutations that block the release of these factors prevent the nucleolar exit of pre-ribosomes. To more specifically assess the roles of these IDRs in ribosome assembly as well as ribosome flux through the nucleolar and nuclear compartments, we deleted the non-visible IDRs in select AFs and assessed growth, polysome profiles and pre-ribosome composition. Certain IDR deletions led to slow growth or temperature-sensitive phenotypes, a decrease in 60S subunits, altered composition of pre-ribosomes and sensitivity to the mistranslation-inducing antibiotic paromomycin. Similarly to IDRs, non-compacted rRNA is also often poorly resolved in early pre-ribosomal intermediates. Hence, it is possible that this non-compacted rRNA is involved in trans interactions between pre-ribosomal particles. Thus, we also want to determine the contribution of rRNA to nucleolar phase separation and release of pre-ribosomes from the nucleolar condensate.
Keywords: ribosome assembly, nucleolus, intrinsically disordered