2013 Rustbelt RNA Meeting







Talk abstracts

Talk on Friday 02:00-02:15pm submitted by Michael Gamalinda

A hierarchical view of the eukaryotic large ribosomal subunit assembly line

Michael Gamalinda (Dept. of Biological Sciences, Carnegie Mellon University), Uli Ohmayer (Lehrstuhl fr Biochemie III, Universitt Regensburg), Jelena Jakovljevic, Beril Kumcuoglu (Dept. of Biological Sciences, Carnegie Mellon University), Joshua Woolford, Betty Mbom (Dept. of Biological Sciences, Carnegie Mellon University), Lawrence Lin (Dept. of Biological Sciences, Carnegie Mellon University), John L. Woolford, Jr. (Dept. of Biological Sciences, Carnegie Mellon University)

Despite having high-resolution structures for eukaryotic large ribosomal subunits, it remained unclear how these ribonucleoprotein complexes are constructed in living cells. Nevertheless, knowing where ribosomal proteins interact with rRNA provides a strategic platform to investigate the connection between spatial and temporal aspects of 60S subunit biogenesis. We previously found that the function of individual yeast large subunit ribosomal proteins (RPLs) in pre-rRNA processing correlates with their location in the structure of mature 60S subunits. This observation suggested that there is an order by which 60S subunits are formed. To directly test this model, we used proteomic approaches to characterize RPLs functioning in early, middle, and late steps of pre-60S assembly by proteomic approaches, assaying the levels of assembly factors and other ribosomal proteins in pre-ribosomes after depleting each RPL. Our results demonstrate that structural domains of eukaryotic 60S complexes are formed in a hierarchical fashion. Assembly begins at the convex solvent-side, followed by the polypeptide exit tunnel, the inter-subunit side, and finally the central protuberance. This model provides an initial paradigm for the sequential assembly of eukaryotic large ribosomal subunits. Our results also reveal striking differences and similarities between assembly of bacterial and eukaryotic large ribosomal subunits, providing insights into the evolution of these RNA-protein particles.

Keywords: ribosome assembly, ribosomal proteins, pre-ribosomes