Yeast polypeptide exit tunnel ribosomal proteins L17, L35 and L37 are necessary to recruit late-assembling factors required for 27SB pre-rRNA processing

Gamalinda, Michael; Jakovljevic, Jelena; Babiano, Reyes; Talkish, Jason; Cruz, Jesús de la; Woolford, John L.

doi:10.1093/nar/gks1272

Cited by 49 publications

(62 citation statements)

References 112 publications

(139 reference statements)

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“…More recently, a refined in vivo assembly map was put forward by Chen and Williamson (2013) that largely corresponds to the Nierhaus map (Rohl and Nierhaus 1982). Our collective studies on yeast RPLs (Babiano et al 2012;Jakovljevic et al 2012;Gamalinda et al 2013;Ohmayer et al 2013) show the following differences from bacterial large subunit assembly: First, while incorporation of bacterial RPLs occurs via distinct assembly groups, most yeast RPLs are present in early assembly intermediates but differ in the step of assembly for which they are required and become more stably assembled in a sequential fashion. Second, several assembly relationships observed in bacterial RPLs are not found for their eukaryotic homologs (Supplemental Fig.…”

Section: Discussionmentioning

confidence: 95%

“…We previously found that abortive assembly intermediates are turned over after depleting RPLs functioning in early and middle steps of pre-60S assembly Gamalinda et al 2013). Preribosomes blocked early in 60S assembly are more rapidly degraded than preribosomes blocked at middle steps, indicating that pre-60S complexes are progressively stabilized through-out successive maturation steps.…”

Section: Preribosomes Become More Stable As Assembly Proceedsmentioning

confidence: 99%

“…Knowing these final endpoints of RP localization provides a strategic platform to investigate the connection between spatial and temporal aspects of ribosome biogenesis. Most RPs assemble with preribosomes early in the biogenesis pathway (Kruiswijk et al 1978;Ferreira-Cerca et al 2007;Babiano and de la Cruz 2010;Babiano et al 2012;Gamalinda et al 2013;Ohmayer et al 2013). As the nascent subunits mature, the association of RPs with pre-rRNA is strengthened by dynamic rearrangement of initial protein-RNA encounter complexes (Ferreira-Cerca et al 2007;Adilakshmi et al 2008;Ohmayer et al 2013).…”

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confidence: 99%

“…RP depletions in yeast and siRNA-mediated knockdowns in cultured mammalian cells showed that most RPs are essential for ribosome biogenesis (Ferreira-Cerca et al 2005;Robledo et al 2008;Poll et al 2009;Babiano and de la Cruz 2010;O'Donohue et al 2010;Jakovljevic et al 2012;Gamalinda et al 2013). However, phenotypic analyses of these mutants were primarily limited to assaying effects on pre-rRNA processing and nucleocytoplasmic export of preribosomes.…”

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confidence: 99%

“…The segregation of large subunits into distinct structural domains is not as apparent as for small subunits (Ban et al 2000); the six rRNA domains are more intertwined in large subunits (Holbrook 2008). Hence, it was initially striking to find that depletion of individual yeast large subunit RPs (RPLs) led to specific rather than global pre-rRNA processing defects (Ferreira-Cerca et al 2005;Hofer et al 2007;Robledo et al 2008;Poll et al 2009; Fernandez-Pevida et al 2012;Jakovljevic et al 2012;Gamalinda et al 2013), which are grouped into early, middle, and late classes (Table 1). From these results, we observed a pattern previously not described: RPLs belonging to these phenotypic classes are remarkably clustered into distinct structural neighborhoods of the yeast 60S subunit ( Fig.…”

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confidence: 99%

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A hierarchical model for assembly of eukaryotic 60S ribosomal subunit domains

et al. 2014

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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.

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Section: Discussionmentioning

confidence: 95%

Section: Preribosomes Become More Stable As Assembly Proceedsmentioning

confidence: 99%

mentioning

confidence: 99%

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confidence: 99%

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confidence: 99%

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A hierarchical model for assembly of eukaryotic 60S ribosomal subunit domains

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Processing of preribosomal RNA in Saccharomyces cerevisiae

2014

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Most, if not all RNAs, are transcribed as precursors that require processing to gain functionality. Ribosomal RNAs (rRNA) from all organisms undergo both exo- and endonucleolytic processing. Also, in all organisms, rRNA processing occurs inside large preribosomal particles and is coupled to nucleotide modification, folding of the precursor rRNA (pre-rRNA), and assembly of the ribosomal proteins (r-proteins). In this review, we focus on the processing pathway of pre-rRNAs of cytoplasmic ribosomes in the yeast Saccharomyces cerevisiae, without doubt, the organism where this pathway is best characterized. We summarize the current understanding of the rRNA maturation process, particularly focusing on the pre-rRNA processing sites, the enzymes responsible for the cleavage or trimming reactions and the different mechanisms that monitor and regulate the pathway. Strikingly, the overall order of the various processing steps is reasonably well conserved in eukaryotes, perhaps reflecting common principles for orchestrating the concomitant events of pre-rRNA processing and ribosome assembly.

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Isolation of conditional mutations in genes essential for viability of Cryptococcus neoformans

2016

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Discovering the genes underlying fundamental processes that enable cells to live and reproduce is a technical challenge, because loss of gene function in mutants results in organisms that cannot survive. This study describes a forward genetics method to identify essential genes in fungi, based on the propensity for Agrobacterium tumefaciens to insert T-DNA molecules into the promoters or 5´ untranslated regions of genes and by placing a conditional promoter within the T-DNA. Insertions of the promoter of the GAL7 gene were made in the human pathogen Cryptococcus neoformans. Nine strains of 960 T-DNA insertional mutants screened grew on media containing galactose, but had impaired growth on media containing glucose, which suppresses expression from GAL7. T-DNA insertions were found in the homologs of IDI1, MRPL37, NOC3, NOP56, PRE3 and RPL17, all of which are essential in ascomycete yeasts Saccharomyces cerevisiae or Schizosaccharomyces pombe. Altering the carbon source in the medium provided a system to identify phenotypes in response to stress agents. The pre3 proteosome subunit mutant was further characterized. The T-DNA insertion and phenotype co-segregate in progeny from a cross, and the growth defect is complemented by the reintroduction of the wild type gene into the insertional mutant. A deletion allele was generated in a diploid strain, this heterozygous strain was sporulated, and analysis of the progeny provided additional genetic evidence that PRE3 is essential. The experimental design is applicable to other fungi and has other forward genetic applications such as to isolate over-expression suppressors or enhance the production of traits of interest.

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Yeast polypeptide exit tunnel ribosomal proteins L17, L35 and L37 are necessary to recruit late-assembling factors required for 27SB pre-rRNA processing

Cited by 49 publications

References 112 publications

A hierarchical model for assembly of eukaryotic 60S ribosomal subunit domains

A hierarchical model for assembly of eukaryotic 60S ribosomal subunit domains

Processing of preribosomal RNA in Saccharomyces cerevisiae

Isolation of conditional mutations in genes essential for viability of Cryptococcus neoformans

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