A Protein Inventory of Human Ribosome Biogenesis Reveals an Essential Function of Exportin 5 in 60S Subunit Export

Wild, Thomas; Horváth, Péter; Wyler, Emanuel; Widmann, Barbara; Badertscher, Lukas; Zemp, Ivo; Kozak, Karol; Csúcs, Gábor; Lund, Elsebet; Kutay, Ulrike

doi:10.1371/journal.pbio.1000522

Cited by 192 publications

(229 citation statements)

References 80 publications

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“…Rpl8 was identified in previous screens as a potential ribosome biogenesis factor (Wild et al 2010;Badertscher et al 2015) and could potentially interact with both the 18S and the 28S rRNA by residing close to the intersubunit space (Anger et al 2013). However, the signal intensity for Rpl8 in the 18S rRNA interactome is similar to that in the 28S scrambled control data set (Fig.…”

Section: Determination Of the Rrna Interactomes Of Hela Cells By Specmentioning

confidence: 74%

“…Proteins were classified according to functional roles in ribosome biology, including components of the cytoplasmic ribosome or implication in ribosome biogenesis reported by system-wide screens or other analyses. Three recent ribosomal biogenesis screens were taken into consideration: Wild et al (2010) (siRNA-based biogenesis screen of the 40S and 60S ribosomal subunit using microscopy as a read-out); Tafforeau et al (2013) (siRNA-based rRNA maturation screen using Northern blotting as a read-out); Badertscher et al (2015) (genome-wide siRNA-based biogenesis screen of the 40S ribosomal subunit using microscopy as a read-out). The above-mentioned methods all share the use of streptavidin-biotin interactions to link the oligonucleotides to a support resin in a noncovalent manner.…”

Section: Discussionmentioning

confidence: 99%

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Specific RNP capture with antisense LNA/DNA mixmers

Rogell

Fischer

Rettel

et al. 2017

RNA

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RNA-binding proteins (RBPs) play essential roles in RNA biology, responding to cellular and environmental stimuli to regulate gene expression. Important advances have helped to determine the (near) complete repertoires of cellular RBPs. However, identification of RBPs associated with specific transcripts remains a challenge. Here, we describe "specific ribonucleoprotein (RNP) capture," a versatile method for the determination of the proteins bound to specific transcripts in vitro and in cellular systems. Specific RNP capture uses UV irradiation to covalently stabilize protein-RNA interactions taking place at "zero distance." Proteins bound to the target RNA are captured by hybridization with antisense locked nucleic acid (LNA)/DNA oligonucleotides covalently coupled to a magnetic resin. After stringent washing, interacting proteins are identified by quantitative mass spectrometry. Applied to in vitro extracts, specific RNP capture identifies the RBPs bound to a reporter mRNA containing the Sex-lethal (Sxl) binding motifs, revealing that the Sxl homolog sister of Sex lethal (Ssx) displays similar binding preferences. This method also revealed the repertoire of RBPs binding to 18S or 28S rRNAs in HeLa cells, including previously unknown rRNA-binding proteins.

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Section: Determination Of the Rrna Interactomes Of Hela Cells By Specmentioning

confidence: 74%

Section: Discussionmentioning

confidence: 99%

Specific RNP capture with antisense LNA/DNA mixmers

Rogell

Fischer

Rettel

et al. 2017

RNA

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“…Previous work has documented that free RPs accumulate in the nucleus/nucleolus, whereas their cytosolic location represents primarily the ribosome-bound pool (43). Therefore, the reduced cytosolic/ nuclear ratio of transiently expressed RPs can be used as a sensitive indicator of disrupted ribosome synthesis (43). However, not all fluorescently tagged RPs assemble into ribosomal subunits when overexpressed at particular levels in a particular cell type (44).…”

Section: Ribosome Biogenesis Is Down-regulated During Forebrainmentioning

confidence: 99%

Requirement of Neuronal Ribosome Synthesis for Growth and Maintenance of the Dendritic Tree

Słomnicki

Pietrzak

Vashishta

et al. 2016

Journal of Biological Chemistry

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The nucleolus serves as a principal site of ribosome biogenesis but is also implicated in various non-ribosomal functions, including negative regulation of the pro-apoptotic transcription factor p53. Although disruption of the nucleolus may trigger the p53-dependent neuronal death, neurotoxic consequences of a selective impairment of ribosome production are unclear. Here, we report that in rat forebrain neuronal maturation is associated with a remarkable expansion of ribosomes despite postnatal down-regulation of ribosomal biogenesis. In cultured rat hippocampal neurons, inhibition of the latter process by knockdowns of ribosomal proteins S6, S14, or L4 reduced ribosome content without disrupting nucleolar integrity, cell survival, and signaling responses to the neurotrophin brain-derived neurotrophic factor. Moreover, reduced general protein synthesis and/or formation of RNA stress granules suggested diminished ribosome recruitment to at least some mRNAs. Such a translational insufficiency was accompanied by impairment of brain-derived neurotrophic factor-mediated dendritic growth. Finally, RNA stress granules and smaller dendritic trees were also observed when ribosomal proteins were depleted from neurons with established dendrites. Thus, a robust ribosomal apparatus is required to carry out protein synthesis that supports dendritic growth and maintenance. Consequently, deficits of ribosomal biogenesis may disturb neurodevelopment by reducing neuronal connectivity. Finally, as stress granule formation and dendritic loss occur early in neurodegenerative diseases, disrupted homeostasis of ribosomes may initiate and/or amplify neurodegeneration-associated disconnection of neuronal circuitries.The ribosome is the principal component of the protein synthesis machinery. Most steps of ribosomal biogenesis occur in the nucleolus starting with the RNA-polymerase-1 (pol 1) 2 -mediated transcription of rRNA genes (1). In addition, pol 1 activity is required for the maintenance of the nucleolus. Therefore, inhibition of pol 1 not only blocks ribosomal biogenesis but also perturbs non-ribosomal functions of the nucleolus, including negative regulation of the pro-apoptotic transcription factor p53 (2). Although pol 1 initiates ribosome production, many additional steps are required for that process, including rRNA processing and binding to ribosomal proteins (RPs) (3). Consequently, depletion of various RPs, including S6 or S14, disrupts ribosomal biogenesis without interfering with nucleolar integrity (4). Impaired ribosome production and/or ribosomal deficits are documented in various neurodegenerative diseases, including Alzheimer, Huntington, Parkinson, amyotrophic lateral sclerosis (ALS), and fronto-temporal lobe dementia (FTLD) (5-13). Reduced ribosomal biogenesis may also contribute to neurodevelopmental disorders. Thus, microcephaly and neurodevelopmental delays are present in Bowen-Conradi syndrome that is caused by deficiency of the ribosomal biogenesis factor EMG1 or in patients with a newly described RPL10 ribos...

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“…28 Remarkably, also Exportin-5 has been shown to export 60S subunit in a Ran-GTP-dependent manner, indicating that robust and overlapping pathways contribute to ribosome export. 29 Accumulating evidence suggests that CRM1 is involved in transport between intranuclear structures and has functions unrelated to nuclear export. 30,31 CRM1 affects shuttling of the U3 snoRNA from Cajal bodies to the nucleolus, 30 binds snoRNA assembly complexes 32 and influences the composition of the nucleoplasmic pre-snoRNP complexes.…”

Section: Crm1mentioning

confidence: 99%

CRM1 and its ribosome export adaptor NMD3 localize to the nucleolus and affect rRNA synthesis

Bai

Moore

Laiho

2013

Nucleus

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A Protein Inventory of Human Ribosome Biogenesis Reveals an Essential Function of Exportin 5 in 60S Subunit Export

Abstract: A systematic search for human ribosome biogenesis factors shows conservation of many aspects of eukaryotic ribosome synthesis with the well-studied process in yeast and identifies an export route of 60S subunits that is specific for higher eukaryotes.

Cited by 192 publications

References 80 publications

Specific RNP capture with antisense LNA/DNA mixmers

Specific RNP capture with antisense LNA/DNA mixmers

Requirement of Neuronal Ribosome Synthesis for Growth and Maintenance of the Dendritic Tree

CRM1 and its ribosome export adaptor NMD3 localize to the nucleolus and affect rRNA synthesis

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