When Good Ribosomes Go Bad

Abstract

Ribosome biogenesis, beyond being a critical requirement for growth in all eukaryotic cells, has a broad impact on the etiology of numerous human disorders. Human genetic diseases caused by mutations in structural components of the ribosome or in factors required to make ribosomes in the cell nucleolus are called ribosomopathies. The congenital ribosomopathy, ANE syndrome, is caused by a single amino acid substitution (L351P) in the third RNA recognition motif (RRM) of the nucleolar protein, RBM28. While patients with ANE syndrome have reduced numbers of mature ribosomes, it is unclear how this single point mutation disrupts nucleolar function in ribosome synthesis and assembly. To investigate the molecular basis of ANE syndrome, we utilized yeast as a model system and determined how the orthologous mutation (L306P) in RRM3 of Nop4, the yeast ortholog of RBM28, contributes to defective ribosome biogenesis. We find that introduction of the ANE syndrome mutation into Nop4 confers both growth and pre‐rRNA processing defects in yeast. Recently, we showed that Nop4 is a hub protein in the nucleolar large subunit (LSU) processome interactome (Genes & Dev. 2015). Here we demonstrate that the presence of the L306P mutation abrogates several, but not all, of Nop4's protein‐protein interactions, thereby disrupting Nop4's hub protein function. Surprisingly, RRMs 3 and 4 of Nop4 are necessary and sufficient for growth, pre‐LSU rRNA processing and hub protein function. Circular dichroism and NMR demonstrate that introduction of the ANE syndrome mutation into RRM3 of human RBM28 disrupts domain folding, consistent with the nucleolar dysfunction of the mutated full‐length protein. We conclude that the ANE syndrome mutation generates defective protein folding which, in turn, abrogates protein‐protein interactions and causes defective pre‐LSU rRNA processing, thus revealing the underlying molecular basis of this human disease.Support or Funding InformationNIGMS, NIEHS, Intramural Research Program of the NIH