Ltv1 Is Required for Efficient Nuclear Export of the Ribosomal Small Subunit in <i>Saccharomyces cerevisiae</i>

Seiser, Robert M.; Sundberg, Alexandra E.; Wollam, Bethany J.; Zobel-Thropp, Pamela A.; Baldwin, Katherine; Spector, Maxwell D.; Lycan, D E

doi:10.1534/genetics.106.062117

Cited by 79 publications

(115 citation statements)

References 55 publications

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“…Of these, human Rio2 has been shown to have a sequence in its C terminus, deletion of which significantly reduces the rate of pre-40S export in human cells (Zemp et al 2009). In yeast, both Dim2 and Ltv1 have been proposed as pre-40S adapters (Seiser et al 2006;Vanrobays et al 2008). However, the proposed NES in Dim2 does not match the new structure-based LR-NES consensus (Guttler et al 2010), and a possible NES in Ltv1 (aa 169-176) was later shown by us to be important for cytoplasmic maturation, but not export (Fassio et al 2010).…”

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confidence: 97%

Genetic Analysis of the Ribosome Biogenesis Factor Ltv1 ofSaccharomyces cerevisiae

et al. 2014

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Ribosome biogenesis has been studied extensively in the yeast Saccharomyces cerevisiae. Yeast Ltv1 is a conserved 40S-associated biogenesis factor that has been proposed to function in small subunit nuclear export. Here we show that Ltv1 has a canonical leucine-rich nuclear export signal (NES) at its extreme C terminus that is both necessary for Crm1 interaction and Ltv1 export. The C terminus of Ltv1 can substitute for the NES in the 60S-export adapter Nmd3, demonstrating that it is a functional NES. Overexpression of an Ltv1 lacking its NES (Ltv1ΔC13) was strongly dominant negative and resulted in the nuclear accumulation of RpS3-GFP; however, export of the pre-40S was not affected. In addition, expression of endogenous levels of Ltv1ΔC protein complemented both the slow-growth phenotype and the 40S biogenesis defect of an ltv1 deletion mutant. Thus, if Ltv1 is a nuclear export adapter for the pre-40S subunit, its function must be fully redundant with additional export factors. The dominant negative phenotype of Ltv1ΔNES overexpression was suppressed by co-overexpressing RpS3 and its chaperone, Yar1, or by deletion of the RpS3-binding site in Ltv1ΔNES, suggesting that titration of RpS3 by Ltv1ΔNES is deleterious in yeast. The dominant-negative phenotype did not correlate with a decrease in 40S levels but rather with a reduction in the polysome-to-monosome ratio, indicating reduced rates of translation. We suggest that titration of RpS3 by excess nuclear Ltv1 interferes with 40S function or with a nonribosomal function of RpS3.

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confidence: 97%

Genetic Analysis of the Ribosome Biogenesis Factor Ltv1 ofSaccharomyces cerevisiae

et al. 2014

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“…The factors involved in 40 S export are largely unknown, but the non-essential pre-40 S factor Ltv1 was suggested to play a role in this process (6). Following nuclear export, the pre-40 S subunit undergoes cytoplasmic maturation, involving two major events.…”

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confidence: 99%

RNA Helicase Prp43 and Its Co-factor Pfa1 Promote 20 to 18 S rRNA Processing Catalyzed by the Endonuclease Nob1

Pertschy

Schneider²,

Gnädig³

et al. 2009

Journal of Biological Chemistry

176

287

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Many RNA nucleases and helicases participate in ribosome biogenesis, but how they cooperate with each other is largely unknown. Here we report that in vivo cleavage of the yeast pre-rRNA at site D, the 3-end of the 18 S rRNA, requires functional interactions between PIN (PilT N terminus) domain protein Nob1 and the DEAH box RNA helicase Prp43. Nob1 showed specific cleavage on a D-site substrate analogue in vitro, which was abolished by mutations in the Nob1 PIN domain or the RNA substrate. Genetic analyses linked Nob1 to the late pre-40 S-associated factor Ltv1, the RNA helicase Prp43, and its cofactor Pfa1. In strains lacking Ltv1, mutation of Prp43 or Pfa1 led to a striking accumulation of 20 S pre-rRNA in the cytoplasm due to inhibition of site D cleavage. This phenotype was suppressed by increased dosage of wildtype Nob1 but not by Nob1 variants mutated in the catalytic site. In ltv1/pfa1 mutants the 20 S pre-rRNA was susceptible to 3 to 5 degradation by the cytoplasmic exosome. This degraded into the 3 region of the 18 S rRNA, strongly indicating that the preribosomes are structurally defective.Eukaryotic ribosome formation is a complex process that requires coordination of rRNA synthesis and processing with the subsequent incorporation of the ribosomal (Rpl and Rps) proteins (reviewed in Refs. 1-5). In the yeast Saccharomyces cerevisiae, ribosome synthesis starts with the transcription of a 35 S pre-rRNA, which is the common precursor to the mature 18, 5.8, and 25 S rRNAs. This pre-rRNA is co-transcriptionally bound by ribosomal proteins as well as many non-ribosomal trans-acting factors to form a 90 S preribosomal particle. Next, this 90 S intermediate undergoes a series of early pre-rRNA processing and modification steps in the nucleolus before an endonucleolytic cleavage at site A 2 in the pre-rRNA separates the pathways for 60 and 40 S subunit assembly.Pre-60 S particles undergo further maturation steps in the nucleus, whereas pre-40 S particles contain few non-ribosomal proteins and are rapidly exported to the cytoplasm. The factors involved in 40 S export are largely unknown, but the non-essential pre-40 S factor Ltv1 was suggested to play a role in this process (6). Following nuclear export, the pre-40 S subunit undergoes cytoplasmic maturation, involving two major events. (i) 20 S pre-rRNA cleavage at site D generates the 3Ј-end of the mature 18 S rRNA, and (ii) structural reorganization forms the characteristic beak structure of the mature 40 S subunit, resulting in exposure of RNA helix 33 within the 18 S rRNA (7).The sites and the order of endonuclease and exonuclease processing events that convert the 35 S pre-rRNA into the mature rRNA species are well characterized in yeast (Fig. 3A) (reviewed in Refs. 4 and 8). However, although most or all of the exonucleases participating in rRNA processing are known, several predicted endonucleases remain to be identified. Two proteins, Fap7 and Nob1, were proposed to cleave site D (9 -11), but for neither was endonuclease activity shown. Fap7 is a putativ...

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“…These proteins are involved in a variety of cellular pro- cesses such as membrane cytoskeleton assembly (Sla2 [32]), mitotic spindle assembly (Spc72 [35]), and chromatin modification (Hpa3 [77]). For Ltv1, a role in the Xpo1-mediated export of small ribosomal subunits has recently been shown (59). No function has been assigned to Sgm1 so far, and with the exception of Ltv1, none of the proteins found in our screen has been analyzed with respect to a possible function in nuclear export.…”

Section: Resultsmentioning

confidence: 92%

Nuclear Export Receptor Xpo1/Crm1 Is Physically and Functionally Linked to the Spindle Pole Body in Budding Yeast

Neuber

Franke

Wittstruck

et al. 2008

Molecular and Cellular Biology

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The spindle pole body (SPB) represents the microtubule organizing center in the budding yeast Saccharomyces cerevisiae. It is a highly structured organelle embedded in the nuclear membrane, which is required to anchor microtubules on both sides of the nuclear envelope. The protein Spc72, a component of the SPB, is located at the cytoplasmic face of this organelle and serves as a receptor for the ␥-tubulin complex. In this paper we show that it is also a binding partner of the nuclear export receptor Xpo1/Crm1. Xpo1 binds its cargoes in a Ran-dependent fashion via a short leucine-rich nuclear export signal (NES). We show that binding of Spc72 to Xpo1 depends on Ran-GTP and a functional NES in Spc72. Mutations in this NES have severe consequences for mitotic spindle morphology in vivo. This is also the case for xpo1 mutants, which show a reduction in cytoplasmic microtubules. In addition, we find a subpopulation of Xpo1 localized at the SPB. Based on these data, we propose a functional link between Xpo1 and the SPB and discuss a role for this exportin in spindle biogenesis in budding yeast.In eukaryotes, the movement of proteins and RNAs between the nucleus and the cytoplasm is an essential cellular process that is mediated by soluble transport receptors (26,52). In addition, the small GTPase Ran (Gsp1 in yeast) and its cytoplasmic and nuclear effectors are needed (12). Transport receptors involved in nuclear import reactions are usually termed importins, and they recognize their cargoes via a specific amino acid motif, the nuclear localization sequence, or NLS. By analogy, nuclear export reactions are mediated by so-called exportins. Xpo1 (also known as Crm1) was one of the first nuclear export receptors to be identified (23,27,50,64) and recognizes a short leucine-rich amino acid sequence on its cargo, called a nuclear export signal, or NES. Numerous export cargoes have now been identified for the mammalian Crm1 protein, including protein kinase A inhibitor PKI, p53, and the human immunodeficiency virus protein Rev (20,25,66). In addition, Crm1 is involved in the nuclear export of several classes of RNAs such as viral pre-mRNAs, ribosomal RNAs, and snRNAs (54). In contrast, few NES-containing proteins have been identified in Saccharomyces cerevisiae. Although the budding yeast Crm1 orthologue Xpo1 is involved in nuclear export of Hog1 (19), Yap1 (76), Ssb1 (62), Ace2 (36), and Prp40 (46), few of the NESs involved have been characterized at a molecular level.Recently, alongside its well-established role in nuclear export, mammalian Crm1 was shown to be involved in the control of centrosome duplication (71). Centrosomes function as microtubule (MT) organizing centers in mammalian somatic cells and direct the formation of a bipolar spindle during mitosis (reviewed in reference 6). In another study, Crm1 was found to bind to kinetochores, complex protein assemblies that are needed to establish the attachment of MTs to chromatin (4). These studies point to an unanticipated role of the exportin Crm1 in control of spin...

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Ltv1 Is Required for Efficient Nuclear Export of the Ribosomal Small Subunit in Saccharomyces cerevisiae

Cited by 79 publications

References 55 publications

Genetic Analysis of the Ribosome Biogenesis Factor Ltv1 ofSaccharomyces cerevisiae

Genetic Analysis of the Ribosome Biogenesis Factor Ltv1 ofSaccharomyces cerevisiae

RNA Helicase Prp43 and Its Co-factor Pfa1 Promote 20 to 18 S rRNA Processing Catalyzed by the Endonuclease Nob1

Nuclear Export Receptor Xpo1/Crm1 Is Physically and Functionally Linked to the Spindle Pole Body in Budding Yeast

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