A synthetic peptide bearing the HIV-1 integrase 161-173 amino acid residues mediates active nuclear import and binding to importin alpha: characterization of a functional nuclear localization signal.
In spite of recent efforts to elucidate the nuclear import pathway of the human immunodeficiency virus type 1 (HIV-1) integrase protein (IN), its exact route as well as the domains that mediate its import are still unknown. Here, we show that a synthetic peptide bearing the amino acid residues 161-173 of the HIV-1 IN is able to mediate active import of covalently attached bovine serum albumin molecules into nuclei of permeabilized cells and therefore was designated as nuclear localization signal-IN (NLS(IN)). A peptide bearing residues 161-173 in the reversed order showed low karyophilic properties. Active nuclear import was demonstrated by using fluorescence microscopy and a quantitative ELISA-based assay system. Nuclear import was blocked by addition of the NLS(IN) peptide, as well as by a peptide bearing the NLS of the simian virus 40 T-antigen (NLS-SV40). The NLS(IN) peptide partially inhibited nuclear import mediated by the full-length recombinant HIV-1 IN protein, indicating that the sequence of the NLS(IN) is involved in mediating nuclear import of the IN protein. The NLS(IN) as well as the full-length IN protein interacted specifically with importin alpha, binding of which was blocked by the NLS(IN) peptide itself as well as by the NLS-SV40.
Armon-Omer
Ayelet
A
Department of Biological Chemistry, The Alexander Silberman Institute of Life Sciences, The Hebrew University of Jerusalem, 91904 Jerusalem, Israel.
Graessmann
Adolf
A
Loyter
Abraham
A
D021581:Active Transport, Cell Nucleus / Q000187:drug effects; D000595:Amino Acid Sequence; D019427:HIV Integrase / Q000737:chemistry / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D028884:Karyopherins / Q000378:metabolism*; D008856:Microscopy, Fluorescence; D019913:Nuclear Localization Signals; D010446:Peptide Fragments / Q000138:chemical synthesis / Q000494:pharmacology*; D011485:Protein Binding / Q000187:drug effects; D021381:Protein Transport; D011994:Recombinant Proteins / Q000378:metabolism
D028884:Karyopherins; D019913:Nuclear Localization Signals; D010446:Peptide Fragments; D011994:Recombinant Proteins; D019427:HIV Integrase