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

Neuber, Anja; Franke, Jacqueline; Wittstruck, Angelika; Schlenstedt, Gabriel; Sommer, Thomas; Stade, Katrin

doi:10.1128/mcb.02043-07

Cited by 15 publications

(13 citation statements)

References 75 publications

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“…This interpretation implies that there is either another NES sequence in Ltv1 or that Ltv1 does not function directly in export. We favor the former of these explanations, however, since Ltv1 has twice been reported to interact with Crm1 in a twohybrid assay, once in a genome-wide two-hybrid screen (Ito et al 2001) and more recently in a directed screen for Crm1-interacting proteins using Crm1 as the bait (Neuber et al 2008).…”

Section: Resultsmentioning

confidence: 99%

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Dominant Mutations in the Late 40S Biogenesis Factor Ltv1 Affect Cytoplasmic Maturation of the Small Ribosomal Subunit in Saccharomyces cerevisiae

et al. 2010

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In eukaryotes, 40S and 60S ribosomal subunits are assembled in the nucleus from rRNAs and ribosomal proteins, exported as premature complexes, and processed in final maturation steps in the cytoplasm. Ltv1 is a conserved 40S ribosome biogenesis factor that interacts with pre-40S complexes in vivo and is proposed to function in yeast in nuclear export. Cells lacking LTV1 grow slowly and are significantly impaired in mature 40S subunit production. Here we show that mutation or deletion of a putative nuclear export sequence in LTV1 is strongly dominant negative, but the protein does not accumulate in the nucleus, as expected for a mutation affecting export. In fact, most of the mutant protein is cytoplasmic and associated with pre-40S subunits. Cells expressing mutant Ltv1 have a 40S biogenesis defect, accumulate 20S rRNA in the cytoplasm as detected by FISH, and retain the late-acting biogenesis factor Tsr1 in the cytoplasm. Finally, overexpression of mutant Ltv1 is associated with nuclear retention of 40S subunit marker proteins, RpS2-GFP and RpS3-GFP. We suggest that the proximal consequence of these LTV1 mutations is inhibition of the cytoplasmic maturation of 40S subunits and that nuclear retention of pre-40S subunits is a downstream consequence of the failure to release and recycle critical factors back to the nucleus. R IBOSOME biogenesis is a major biosynthetic activity of eukaryotic cells and a significant ratelimiting factor in cell growth and proliferation. The pathway appears to be conserved in most respects in eukaryotes from yeast to humans and has been extensively analyzed in the model organism, Saccharomyces cerevisiae. Ribosome biogenesis begins cotranscriptionally in the nucleolus, continues in the nucleoplasm, and is completed in the cytoplasm where final maturation events must occur (for reviews, see More than 150 trans-acting factors have been identified by genetic and proteomic studies with roles in ribosome biogenesis. Generally, these factors are well conserved through eukaryotic evolution, but the specific function of many of the proteins remains largely unknown.In yeast, the 18S, 5.8S, and 25S ribosomal RNAs (rRNAs) are transcribed as a single 35S precursor rRNA. About 40 ribosome biogenesis factors, U3 snoRNA, and ribosomal proteins assemble cotranscriptionally on the nascent 35S pre-rRNA to form the 90S pre-ribosome (Dragon et al. 2002;Grandi et al. 2002;Schafer et al. 2003). Cleavage of the 35S pre-rRNA at site A2 releases a pre-40S particle containing 20S pre-rRNA, which then sheds most of the processing factors associated with it (Schafer et al. 2003). Pre-60S biogenesis factors and ribosomal proteins then assemble on the remaining transcript. The pre-60S subunit undergoes extensive remodeling and processing in the nucleolus and nucleoplasm, acquiring new processing factors and incorporating the independently transcribed 5S pre-rRNA before being exported through the nuclear pore to the cytoplasm (reviewed in Fromont-Racine et al. 2003;Tschochner and Hurt 2003). The pre-40S an...

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Section: Resultsmentioning

confidence: 99%

“…(Seiser et al 2006). Ltv1 also interacts with Crm1 in a yeast two-hybrid assay (Ito et al 2001;Neuber et al 2008;our unpublished results).…”

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confidence: 99%

Dominant Mutations in the Late 40S Biogenesis Factor Ltv1 Affect Cytoplasmic Maturation of the Small Ribosomal Subunit in Saccharomyces cerevisiae

et al. 2010

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“…CRM1 has also been identified at centrosomes in a number of model systems and its interaction with specific NES-containing proteins can regulate their impact on centrosome duplication (43,55,56). CRM1 NES binding is required for the proper localization of centrosomal proteins centrin and pericentrin (57) and breast cancer 2 susceptibility protein (BRCA2) (58,59).…”

Section: Discussionmentioning

confidence: 99%

The huntingtin N17 domain is a multifunctional CRM1 and Ran-dependent nuclear and cilial export signal

Maiuri

Woloshansky

Xia

et al. 2013

Human Molecular Genetics

116

136

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The first 17 amino acids of Huntington’s disease (HD) protein, huntingtin, comprise an amphipathic alpha-helical domain that can target huntingtin to the endoplasmic reticulum (ER). N17 is phosphorylated at two serines, shown to be important for disease development in genetic mouse models, and shown to be modified by agents that reverse the disease phenotype in an HD mouse model. Here, we show that the hydrophobic face of N17 comprises a consensus CRM1/exportin-dependent nuclear export signal, and that this nuclear export activity can be affected by serine phospho-mimetic mutants. We define the precise residues that comprise this nuclear export sequence (NES) as well as the interaction of the NES, but not phospho-mimetic mutants, with the CRM1 nuclear export factor. We show that the nuclear localization of huntingtin depends upon the RanGTP/GDP gradient, and that N17 phosphorylation can also distinguish localization of endogenous huntingtin between the basal body and stalk of the primary cilium. We present a mechanism and multifunctional role for N17 in which phosphorylation of N17 not only releases huntingtin from the ER to allow nuclear entry, but also prevents nuclear export during a transient stress response event to increase the levels of nuclear huntingtin and to regulate huntingtin access to the primary cilium. Thus, N17 is a master localization signal of huntingtin that can mediate huntingtin localization between the cytoplasm, nucleus and primary cilium. This localization can be regulated by signaling, and is misregulated in HD.

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“…However, 42 (16%) of the NESs in ValidNESs have not had their CRM1 dependence validated with LMB. For these NESs, some other experimental techniques, such as yeast two-hybrid system and in vitro binding experiments, were used to demonstrate the interaction between CRM1- and NES-containing proteins (17,18). However, many of these NESs, 27 from NESbase for instance, were discovered around the early 2000s.…”

Section: Data Curationmentioning

confidence: 99%

ValidNESs: a database of validated leucine-rich nuclear export signals

Fu¹,

Huang²,

Horton³

et al. 2012

Nucleic Acids Research

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ValidNESs (http://validness.ym.edu.tw/) is a new database for experimentally validated leucine-rich nuclear export signal (NES)-containing proteins. The therapeutic potential of the chromosomal region maintenance 1 (CRM1)-mediated nuclear export pathway and disease relevance of its cargo proteins has gained recognition in recent years. Unfortunately, only about one-third of known CRM1 cargo proteins are accessible in a single database since the last compilation in 2003. CRM1 cargo proteins are often recognized by a classical NES (leucine-rich NES), but this signal is notoriously difficult to predict from sequence alone. Fortunately, a recently developed prediction method, NESsential, is able to identify good candidates in some cases, enabling valuable hints to be gained by in silico prediction, but until now it has not been available through a web interface. We present ValidNESs, an integrated, up-to-date database holding 221 NES-containing proteins, combined with a web interface to prediction by NESsential.

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

Cited by 15 publications

References 75 publications

Dominant Mutations in the Late 40S Biogenesis Factor Ltv1 Affect Cytoplasmic Maturation of the Small Ribosomal Subunit in Saccharomyces cerevisiae

Dominant Mutations in the Late 40S Biogenesis Factor Ltv1 Affect Cytoplasmic Maturation of the Small Ribosomal Subunit in Saccharomyces cerevisiae

The huntingtin N17 domain is a multifunctional CRM1 and Ran-dependent nuclear and cilial export signal

ValidNESs: a database of validated leucine-rich nuclear export signals

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