Eukaryotic Ribosome assembly and Nucleocytoplasmic Transport

Abstract

The process of eukaryotic ribosome assembly stretches across the nucleolus, the nucleoplasm and the cytoplasm, and therefore relies on efficient nucleocytoplasmic transport. In yeast, the import machinery delivers ~140,000 ribosomal proteins every minute to the nucleus for ribosome assembly. At the same time, the export machinery facilitates translocation of ~2000 pre-ribosomal particles every minute through ~200 nuclear pore complexes (NPC) into the cytoplasm. Eukaryotic ribosome assembly also requires >200 conserved assembly factors, which transiently associate with pre-ribosomal particles. Their site(s) of action on maturing pre-ribosomes are beginning to be elucidated. In this chapter, we outline protocols that enable rapid biochemical isolation of pre-ribosomal particles for single particle cryo-electron microscopy (cryo-EM) and in vitro reconstitution of nuclear transport processes. We discuss cell-biological and genetic approaches to investigate how the ribosome assembly and the nucleocytoplasmic transport machineries collaborate to produce functional ribosomes.

Ribosomal Biogenesis and Heterogeneity in Development, Disease, and Aging

“…Clusters of 60–800 copies of 45S precursor rDNA genes encoding 5.8S, 18S, and 28S rRNAs are located in the acrocentric chromosomes (chromosomes 13, 14, 15, 21, and 22 in humans) in nucleolus organiser regions (NORs). There are 10–400 copies of 5S rRNA located on chromosome 1; 45S/35S rDNAs are transcribed by RNA Pol I in the nucleolus, and 5S rDNA is transcribed by RNA Pol III generally in the nucleoplasm, with an exception in yeast, where it is likely to work in the nucleolus due to 5S rDNA’s interspersed location with 35S rDNA on chromosome XII [ 8 , 9 , 10 ]. RNA Pol II transcribes all ribosomal-protein-coding genes in the nucleoplasm.…”

“…The mRNAs of ribosomal proteins (RPs) translocate to the cytoplasm and get translated, and, in most cases, RPs return to the nucleus. The precursor subunits are assembled in the nucleolus and in the nucleoplasm before being transported to the cytoplasm where they undergo further modifications to generate the mature and functional ribosomes [ 8 , 9 ]. Figure 1 illustrates the ribosome biogenesis process in yeast.…”

“…transcribed by RNA Pol III generally in the nucleoplasm, with an exception in yeast, where it is likely to work in the nucleolus due to 5S rDNA's interspersed location with 35S rDNA on chromosome XII [8][9][10]. RNA Pol II transcribes all ribosomal-protein-coding genes in the nucleoplasm.…”

Inhibition of Ribosome Assembly and Ribosome Translation Has Distinctly Different Effects on Abundance and Paralogue Composition of Ribosomal Protein mRNAs in Saccharomyces cerevisiae

“…Mutations in numerous genes for r-proteins or assembly factors cause a variety of congenital human diseases, collectively called ribosomopathies ( 14 – 17 ). It is assumed that the root for these calamities is the decreased translation capacity, which in turn can lead to the formation of specialized ribosomes with altered preferences for specific mRNAs, the interaction of free r-proteins with p53 and cell cycle regulators, and changes to specific metabolic pathways ( 18 – 23 ).…”

Inhibiting ribosome assembly and ribosome translation have distinctly different effects on the abundance and paralogue composition of ribosomal protein mRNAs inSaccharomyces cerevisiae

“…Mutations in numerous genes for r-proteins or assembly factors cause a variety of congenital human diseases, collectively called "ribosomopathies" (14)(15)(16)(17). It is assumed that the root for these calamities is the decreased translation capacity, which in turn can lead to the formation of "specialized ribosomes" with altered preferences for specific mRNAs, the interaction of free r-proteins with p53 and cell cycle regulators, and changes to specific metabolic pathways (18)(19)(20)(21)(22)(23).…”