A conserved phosphoprotein that specifically binds nuclear localization sequences is involved in nuclear import.
We have purified proteins of 70 kD from Drosophila, HeLa cells, and Z. mays that specifically bind nuclear localization sequences (NLSs). These proteins are recognized by antibodies raised against a previously identified NLS-binding protein (NBP) from the yeast S. cerevisiae. All NBPs are associated with nuclei and also present in the cytosol. NBPs are phosphorylated and phosphatase treatment abolished NLS binding. The requirement for NBPs in nuclear protein uptake is demonstrated in semipermeabilized Drosophila melanogaster tissue culture cells. Proper import of a fluorescent protein containing the large T antigen NLS requires cytosol and ATP. In the absence of cytosol and/or ATP, NLS-containing proteins are bound to cytosolic structures and the nuclear envelope. Addition of cytosol and ATP results in movement of this bound intermediate into the nucleus. Anti-NBP antibodies specifically inhibited the binding part of this import reaction. These results indicate that a phosphoprotein common to several eukaryotes acts as a receptor that recognizes NLSs before their uptake into the nucleus.
Stochaj
U
U
Department of Molecular Biology, Princeton University, New Jersey 08544.
Silver
P A
PA
D000818:Animals; D001693:Biological Transport, Active; D002467:Cell Nucleus / Q000737:chemistry / Q000378:metabolism*; D002478:Cells, Cultured; D003593:Cytoplasm / Q000737:chemistry / Q000378:metabolism*; D004330:Drosophila; D005057:Eukaryotic Cells / Q000737:chemistry / Q000378:metabolism; D005455:Fluorescent Antibody Technique; D006367:HeLa Cells; D006801:Humans; D019913:Nuclear Localization Signals; D009687:Nuclear Proteins / Q000276:immunology / Q000378:metabolism*; D010750:Phosphoproteins / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism*; D021382:Protein Sorting Signals / Q000276:immunology / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000737:chemistry / Q000378:metabolism; D003313:Zea mays
D019913:Nuclear Localization Signals; D009687:Nuclear Proteins; D010750:Phosphoproteins; D021382:Protein Sorting Signals