Proteins destined for import into the nucleus contain nuclear localization signals (NLSs) that are recognized by import receptors termed karyopherins or importins. Until recently, the only nuclear import sequence that had been well-defined and characterized was the classical NLS (cNLS), which is recognized by importin a. However, Chook and colleagues (Lee et al., 2006, Cell 126, 543-558) have provided new insight into nuclear targeting with their identification of a novel NLS, termed the PY-NLS, that is recognized by the human karyopherin ss2/transportin (Kapss2) receptor. Here, we demonstrate that the PY-NLS is conserved in S. cerevisiae and show for the first time that the PY-NLS is a functional nuclear targeting sequence in vivo. The apparent ortholog of Kapss2 in yeast, Kap104, has two known cargos, the mRNA-binding proteins Hrp1 and Nab2, which both contain putative PY-NLS-like sequences. We find that the PY-NLS-like sequence within Hrp1, which closely matches the PY-NLS consensus, is both necessary and sufficient for nuclear import and is also required for receptor binding and protein function. In contrast, the PY-NLS-like sequences in Nab2, which vary from the PY-NLS consensus, are not required for proper import or protein function, suggesting that Kap104 may interact with different cargos using multiple mechanisms. Dissection of the PY-NLS consensus reveals that the minimal PY-NLS in yeast consists of the C-terminal portion of the human consensus, R/H/KX2-5PY, with upstream basic or hydrophobic residues enhancing the targeting function. Finally, we apply this analysis to a bioinformatic search of the yeast proteome as a preliminary search for new potential Kap104 cargos.
|Evidence ID||Analyze ID||Interactor||Interactor Systematic Name||Interactor||Interactor Systematic Name||Type||Assay||Annotation||Action||Modification||Phenotype||Source||Reference||Note|
|Evidence ID||Analyze ID||Gene||Gene Systematic Name||Gene Ontology Term||Gene Ontology Term ID||Qualifier||Aspect||Method||Evidence||Source||Assigned On||Annotation Extension||Reference|
|Evidence ID||Analyze ID||Gene||Gene Systematic Name||Phenotype||Experiment Type||Experiment Type Category||Mutant Information||Strain Background||Chemical||Details||Reference|
|Evidence ID||Analyze ID||Regulator||Regulator Systematic Name||Target||Target Systematic Name||Experiment||Assay||Construct||Conditions||Strain Background||Reference|