Identification of a nuclear localization signal of a yeast ribosomal protein.
To identify a signal involved in transporting a ribosomal protein to the nucleus, we constructed hybrid genes encoding amino-terminal segments of yeast ribosomal protein L3 joined to the amino-terminal end of the entire Escherichia coli beta-galactosidase molecule. The subcellular locations of the corresponding hybrid proteins in yeast were determined by in situ immunofluorescence. The first 21 amino acids of L3 were sufficient to localize beta-galactosidase to the nucleus. This region shows limited homology to portions of other nuclear proteins identified as essential for their transport. Larger fusion proteins were also localized to the nucleus. However, a hybrid protein containing all but the 14 carboxyl-terminal amino acids from L3 initially failed to localize; this defect was corrected by inserting a glycine- and proline-containing bridge between the L3 and beta-galactosidase moieties. The renovated protein was able to associate with ribosomes, suggesting that, in addition to entering the nucleus, this hybrid polypeptide was assembled into 60S ribosomal subunits that were subsequently exported to the cytoplasm.
D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D001693:Biological Transport, Active; D002467:Cell Nucleus / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D005455:Fluorescent Antibody Technique; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism*; D011487:Protein Conformation; D021382:Protein Sorting Signals / Q000235:genetics*; D011994:Recombinant Proteins / Q000032:analysis; D000090822:Ribosomal Protein L3; D012269:Ribosomal Proteins / Q000235:genetics / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D012689:Sequence Homology, Nucleic Acid; D001616:beta-Galactosidase / Q000235:genetics / Q000378:metabolism
D001426:Bacterial Proteins; D005656:Fungal Proteins; D021382:Protein Sorting Signals; D011994:Recombinant Proteins; D000090822:Ribosomal Protein L3; D012269:Ribosomal Proteins; D001616:beta-Galactosidase