A hierarchical model for assembly of eukaryotic 60S ribosomal subunit domains.
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 ribosomal RNA (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 precursor rRNA (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 test this model, we used proteomic approaches to assay changes in the levels of ribosomal proteins and assembly factors in preribosomes when RPLs functioning in early, middle, and late steps of pre-60S assembly are depleted. Our results demonstrate that structural domains of eukaryotic 60S ribosomal subunits are formed in a hierarchical fashion. Assembly begins at the convex solvent side, followed by the polypeptide exit tunnel, the intersubunit side, and finally the central protuberance. This model provides an initial paradigm for the sequential assembly of eukaryotic 60S subunits. Our results reveal striking differences and similarities between assembly of bacterial and eukaryotic large ribosomal subunits, providing insights into how these RNA-protein particles evolved.
Department of Biological Sciences, Carnegie Mellon University, Pittsburgh, Pennsylvania 15213, USA;
Michael
M
Gamalinda
Uli
U
Ohmayer
Jelena
J
Jakovljevic
Beril
B
Kumcuoglu
Joshua
J
Woolford
Bertrade
B
Mbom
Lawrence
L
Lin
John L
JL
Woolford
D008958:Models, Molecular; D020836:Protein Structure, Quaternary; D017434:Protein Structure, Tertiary / Q000502:physiology; D054683:Ribosome Subunits, Large, Eukaryotic / Q000737:chemistry / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000737:chemistry / Q000378:metabolism*
large ribosomal subunit; preribosomes; ribosomal proteins; ribosome assembly