Factors affecting nuclear export of the 60S ribosomal subunit in vivo.
In Saccharomyces cerevisiae, the 60S ribosomal subunit assembles in the nucleolus and then is exported to the cytoplasm, where it joins the 40S subunit for translation. Export of the 60S subunit from the nucleus is known to be an energy-dependent and factor-mediated process, but very little is known about the specifics of its transport. To begin to address this problem, an assay was developed to follow the localization of the 60S ribosomal subunit in S. cerevisiae. Ribosomal protein L11b (Rpl11b), one of the approximately 45 ribosomal proteins of the 60S subunit, was tagged at its carboxyl terminus with the green fluorescent protein (GFP) to enable visualization of the 60S subunit in living cells. A panel of mutant yeast strains was screened for their accumulation of Rpl11b-GFP in the nucleus as an indicator of their involvement in ribosome synthesis and/or transport. This panel included conditional alleles of several rRNA-processing factors, nucleoporins, general transport factors, and karyopherins. As predicted, conditional alleles of rRNA-processing factors that affect 60S ribosomal subunit assembly accumulated Rpl11b-GFP in the nucleus. In addition, several of the nucleoporin mutants as well as a few of the karyopherin and transport factor mutants also mislocalized Rpl11b-GFP. In particular, deletion of the previously uncharacterized karyopherin KAP120 caused accumulation of Rpl11b-GFP in the nucleus, whereas ribosomal protein import was not impaired. Together, these data further define the requirements for ribosomal subunit export and suggest a biological function for KAP120.
Stage-Zimmermann
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Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Dana-Farber Cancer Institute, Boston, Massachusetts 02115, USA.
Schmidt
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Silver
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PA
D021581:Active Transport, Cell Nucleus; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism; D002455:Cell Division; D002467:Cell Nucleus / Q000378:metabolism*; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism*; D017353:Gene Deletion; D049452:Green Fluorescent Proteins; D028884:Karyopherins; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D008967:Molecular Biology / Q000379:methods; D009154:Mutation; D028861:Nuclear Pore Complex Proteins; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism; D029543:Nucleocytoplasmic Transport Proteins; D012322:RNA Precursors / Q000378:metabolism; D012323:RNA Processing, Post-Transcriptional; D016601:RNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D018160:Receptors, Cytoplasmic and Nuclear; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012269:Ribosomal Proteins / Q000235:genetics / Q000378:metabolism*; D012270:Ribosomes / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000254:growth & development / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins; D028961:beta Karyopherins; D000097606:Exportin 1 Protein
D002352:Carrier Proteins; D005656:Fungal Proteins; D049452:Green Fluorescent Proteins; D028884:Karyopherins; D008164:Luminescent Proteins; D028861:Nuclear Pore Complex Proteins; D009687:Nuclear Proteins; D029543:Nucleocytoplasmic Transport Proteins; D012322:RNA Precursors; D016601:RNA-Binding Proteins; D018160:Receptors, Cytoplasmic and Nuclear; D011993:Recombinant Fusion Proteins; D012269:Ribosomal Proteins; D029701:Saccharomyces cerevisiae Proteins; D028961:beta Karyopherins; D000097606:Exportin 1 Protein; C082274:CSE1 protein, S cerevisiae; C101820:KAP104 protein, S cerevisiae; C401306:MSN5 protein, S cerevisiae; C110033:MTR10 protein, S cerevisiae; C077734:NPL3 protein, S cerevisiae; C097740:NUP120 protein, S cerevisiae; C027624:ribosomal protein L11