Invariable Ribosome Stoichiometry During Murine Erythroid Differentiation: Implications for Understanding Ribosomopathies

Abstract

Heterogeneity of the main ribosomal composition represents an emerging, yet debatable, mechanism of gene expression regulation with a purported role in ribosomopathies, a group of disorders caused by mutations in ribosomal protein genes (RPs). Ribosomopathies, mysteriously relate with tissue-specific symptoms (mainly anemia and cancer predisposition), despite the ubiquitous expression and necessity of the associated RPs. An outstanding question that may shed light into disease pathogenicity and provide potential pharmacological interventions, is whether and how the ribosomal composition is modified during, the highly affected by RP mutations, process of erythroid differentiation. To address this issue, we analyzed ribosome stoichiometry using an established model of erythroid differentiation, through sucrose gradient ultracentrifugation and quantitative proteomics. We found that differentiation associates with an extensive reprogramming of the overall ribosomal levels, characterized by an increase in monosomes and a decrease in polysomes. However, by calculating a stoichiometry score for each independent ribosomal protein, we found that the main ribosomal architecture remained invariable between immature and differentiated cells. In total, none of the 78 Ribosomal Proteins (RPs- 74 core RPs, Rack1, Fau and 2 paralogs) detected was statistically different between the samples. This data was further verified through antibody-mediated quantification of 6 representative RPs. Moreover, bioinformatic analysis of whole cell proteomic data derived out of 4 additional models of erythropoiesis revealed that RPs were co-regulated across these cell types, too. In conclusion, ribosomes maintain an invariant protein stoichiometry during differentiation, thus excluding ribosome heterogeneity from a potential mechanism of toxicity in ribosomopathies and other erythroid disorders.

HP1 proteins regulate nucleolar structure and function by secluding pericentromeric constitutive heterochromatin

“…Slow growth phenotypes have been reported for several congenital or somatic tissue-specific ribosomopathy diseases which are caused by mutations in ribosomal proteins or ribosome biogenesis factors. Such mutations affect the processing of ribosomal RNAs (rRNAs) and ribonucleoproteins, disrupting the integrity of nucleoli and leading to a shortage of mature ribosomes and generally hypo-proliferation ( 15 , 102 , 103 ).…”

The RNA-Binding Function of Ribosomal Proteins and Ribosome Biogenesis Factors in Human Health and Disease

“…It has also been hypothesized that the tissue-specific character of ribosomopathies may originate from the existence of specialized subgroups of ribosomes with increased affinity for 5 -UTR of transcripts mostly involved in erythroid differentiation. However, stoichiometric analysis of RPs isolated from ribosomes belonging to four cell models of erythropoiesis showed a consistent protein balance throughout the process of cellular differentiation, thus ruling out in this case the possibility of ribosomal diversity as a source of DBA ribosomopathy [139].…”