U3 snoRNA promoter reflects the RNA’s function in ribosome biogenesis

ABSTRACT Small nucleolar RNAs (snoRNAs) orchestrate the modification and cleavage of pre-rRNA and are essential for ribosome biogenesis. Recent data suggest that after nucleoplasmic synthesis, snoRNAs transiently localize to the Cajal body (in plant and animal cells) or the homologous nucleolar body (in budding yeast) for maturation and assembly into snoRNPs prior to accumulation in their primary functional site, the nucleolus. However, little is known about the trans-acting factors important for the intranuclear trafficking and nucleolar localization of snoRNAs. Here, we describe a large-scale genetic screen to identify proteins important for snoRNA transport in Saccharomyces cerevisiae. We performed fluorescence in situ hybridization analysis to visualize U3 snoRNA localization in a collection of temperature-sensitive yeast mutants. We have identified Nop4, Prp21, Tao3, Sec14, and Htl1 as proteins important for the proper localization of U3 snoRNA. Mutations in genes encoding these proteins lead to specific defects in the targeting or retention of the snoRNA to either the nucleolar body or the nucleolus. Additional characterization of the mutants revealed impairment in specific steps of U3 snoRNA processing, demonstrating that snoRNA maturation and trafficking are linked processes.

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Faculty Opinions recommendation of RNA chaperones stimulate formation and yield of the U3 snoRNA-Pre-rRNA duplexes needed for eukaryotic ribosome biogenesis.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.1162632.624200 ◽

2009 ◽

Author(s):

Jonathan R Warner

Keyword(s):

Ribosome Biogenesis ◽

Eukaryotic Ribosome ◽

U3 Snorna ◽

Rna Chaperones

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Role of Pre-rRNA Base Pairing and 80S Complex Formation in Subnucleolar Localization of the U3 snoRNP

Molecular and Cellular Biology ◽

10.1128/mcb.24.19.8600-8610.2004 ◽

2004 ◽

Vol 24 (19) ◽

pp. 8600-8610 ◽

Cited By ~ 33

Author(s):

Sander Granneman ◽

Judith Vogelzangs ◽

Reinhard Lührmann ◽

Walther J. van Venrooij ◽

Ger J. M. Pruijn ◽

...

Keyword(s):

Complex Formation ◽

Ribosome Biogenesis ◽

Specific Protein ◽

Direct Result ◽

Rrna Processing ◽

Base Pairing ◽

Active Involvement ◽

U3 Snorna ◽

Fibrillar Component

ABSTRACT In the nucleolus the U3 snoRNA is recruited to the 80S pre-rRNA processing complex in the dense fibrillar component (DFC). The U3 snoRNA is found throughout the nucleolus and has been proposed to move with the preribosomes to the granular component (GC). In contrast, the localization of other RNAs, such as the U8 snoRNA, is restricted to the DFC. Here we show that the incorporation of the U3 snoRNA into the 80S processing complex is not dependent on pre-rRNA base pairing sequences but requires the B/C motif, a U3-specific protein-binding element. We also show that the binding of Mpp10 to the 80S U3 complex is dependent on sequences within the U3 snoRNA that base pair with the pre-rRNA adjacent to the initial cleavage site. Furthermore, mutations that inhibit 80S complex formation and/or the association of Mpp10 result in retention of the U3 snoRNA in the DFC. From this we propose that the GC localization of the U3 snoRNA is a direct result of its active involvement in the initial steps of ribosome biogenesis.

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Nucleolar proteins Bfr2 and Enp2 interact with DEAD-box RNA helicase Dbp4 in two different complexes

Nucleic Acids Research ◽

10.1093/nar/gkt1293 ◽

2013 ◽

Vol 42 (5) ◽

pp. 3194-3206 ◽

Cited By ~ 16

Author(s):

Sahar Soltanieh ◽

Martin Lapensée ◽

François Dragon

Keyword(s):

Ribosomal Rna ◽

Ribosome Biogenesis ◽

Rna Helicase ◽

Small Subunit ◽

Specific Protein ◽

18S Ribosomal Rna ◽

Dead Box ◽

Nucleolar Proteins ◽

U3 Snorna ◽

Ssu Processome

AbstractDifferent pre-ribosomal complexes are formed during ribosome biogenesis, and the composition of these complexes is highly dynamic. Dbp4, a conserved DEAD-box RNA helicase implicated in ribosome biogenesis, interacts with nucleolar proteins Bfr2 and Enp2. We show that, like Dbp4, Bfr2 and Enp2 are required for the early processing steps leading to the production of 18S ribosomal RNA. We also found that Bfr2 and Enp2 associate with the U3 small nucleolar RNA (snoRNA), the U3-specific protein Mpp10 and various pre-18S ribosomal RNA species. Thus, we propose that Bfr2, Dbp4 and Enp2 are components of the small subunit (SSU) processome, a large complex of ∼80S. Sucrose gradient sedimentation analyses indicated that Dbp4, Bfr2 and Enp2 sediment in a peak of ∼50S and in a peak of ∼80S. Bfr2, Dbp4 and Enp2 associate together in the 50S complex, which does not include the U3 snoRNA; however, they associate with U3 snoRNA in the 80S complex (SSU processome). Immunoprecipitation experiments revealed that U14 snoRNA associates with Dbp4 in the 50S complex, but not with Bfr2 or Enp2. The assembly factor Tsr1 is not part of the ‘50S’ complex, indicating this complex is not a pre-40S ribosome. A combination of experiments leads us to propose that Bfr2, Enp2 and Dbp4 are recruited at late steps during assembly of the SSU processome.

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Utp14 Recruits and Activates the RNA Helicase Dhr1 To Undock U3 snoRNA from the Preribosome

Molecular and Cellular Biology ◽

10.1128/mcb.00773-15 ◽

2016 ◽

Vol 36 (6) ◽

pp. 965-978 ◽

Cited By ~ 29

Author(s):

Jieyi Zhu ◽

Xin Liu ◽

Margarida Anjos ◽

Carl C. Correll ◽

Arlen W. Johnson

Keyword(s):

Ribosome Biogenesis ◽

Rna Helicase ◽

Small Subunit ◽

Genetic Interactions ◽

Base Pairs ◽

Eukaryotic Ribosome ◽

U3 Snorna ◽

Stimulation Of

In eukaryotic ribosome biogenesis, U3 snoRNA base pairs with the pre-rRNA to promote its processing. However, U3 must be removed to allow folding of the central pseudoknot, a key feature of the small subunit. Previously, we showed that the DEAH/RHA RNA helicase Dhr1 dislodges U3 from the pre-rRNA.DHR1can be linked toUTP14, encoding an essential protein of the preribosome, through genetic interactions with the rRNA methyltransferase Bud23. Here, we report that Utp14 regulates Dhr1. Mutations within a discrete region of Utp14 reduced interaction with Dhr1 that correlated with reduced function of Utp14. These mutants accumulated Dhr1 and U3 in a pre-40S particle, mimicking a helicase-inactive Dhr1 mutant. This similarity in the phenotypes led us to propose that Utp14 activates Dhr1. Indeed, Utp14 formed a complex with Dhr1 and stimulated its unwinding activityin vitro. Moreover, theutp14mutants that mimicked a catalytically inactivedhr1mutantin vivoshowed reduced stimulation of unwinding activityin vitro. Dhr1 binding to the preribosome was substantially reduced only when both Utp14 and Bud23 were depleted. Thus, Utp14 is bifunctional; together with Bud23, it is needed for stable interaction of Dhr1 with the preribosome, and Utp14 activates Dhr1 to dislodge U3.

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Inactivation of Cleavage Factor I Components Rna14p and Rna15p Induces Sequestration of Small Nucleolar Ribonucleoproteins at Discrete Sites in the Nucleus

Molecular Biology of the Cell ◽

10.1091/mbc.e07-10-1015 ◽

2008 ◽

Vol 19 (4) ◽

pp. 1499-1508 ◽

Cited By ~ 5

Author(s):

Tiago Carneiro ◽

Célia Carvalho ◽

José Braga ◽

José Rino ◽

Laura Milligan ◽

...

Keyword(s):

Quality Control ◽

Ribosome Biogenesis ◽

Small Nucleolar Rnas ◽

Factor I ◽

U3 Snorna ◽

Specific Proteins ◽

Nuclear Exosome ◽

Nucleolar Rnas

Small nucleolar RNAs (snoRNAs) associate with specific proteins forming small nucleolar ribonucleoprotein (snoRNP) particles, which are essential for ribosome biogenesis. The snoRNAs are transcribed, processed, and assembled in snoRNPs in the nucleoplasm. Mature particles are then transported to the nucleolus. In yeast, 3′-end maturation of snoRNAs involves the activity of Rnt1p endonuclease and cleavage factor IA (CFIA). We report that after inhibition of CFIA components Rna14p and Rna15p, the snoRNP proteins Nop1p, Nop58p, and Gar1p delocalize from the nucleolus and accumulate in discrete nucleoplasmic foci. The U14 snoRNA, but not U3 snoRNA, similarly redistributes from the nucleolus to the nucleoplasmic foci. Simultaneous depletion of either Rna14p or Rna15p and the nuclear exosome component Rrp6p induces accumulation of poly(A)+ RNA at the snoRNP-containing foci. We propose that the foci detected after CFIA inactivation correspond to quality control centers in the nucleoplasm.

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