A ligand-dependent bipartite nuclear targeting signal in the human androgen receptor. Requirement for the DNA-binding domain and modulation by NH2-terminal and carboxyl-terminal sequences.
The amino acid sequence requirements for androgen-dependent androgen receptor nuclear import were determined by immunostaining transiently expressed full-length wild type and mutant human androgen receptors (AR) in monkey kidney COS cells and measuring transcriptional activity by cotransfection with a luciferase reporter vector in monkey kidney CV1 cells. Mutagenesis studies revealed a bipartite nuclear targeting sequence in the DNA binding and hinge regions at amino acids 617-633, consisting of two clusters of basic amino acids separated by 10 amino acids, (sequence: see text). In a series of deletion mutants, AR NH2-terminal fragments (residues 1-639 through 1-723) displayed constitutive nuclear import, and transcriptional activity was similar to that of the ligand-activated full-length wild type AR. In contrast, nuclear import and transcriptional activation were inhibited by sequence extensions into the steroid-binding domain (1-771). Constitutive nuclear import was regained in part by NH2-terminal deletions of full-length AR. Expression of AR/pyruvate kinase chimeras defined a sequence required for pre-dominant nuclear localization as residues 580-661, comprised of the second zinc finger region of the DNA-binding domain, the 17-amino-acid putative targeting sequence, and 28 residues of flanking carboxyl-terminal sequence. These studies suggest that the bipartite nuclear targeting sequence of AR includes flanking sequence and is modulated by interactions between the NH2-and carboxyl-terminal regions.
Zhou
Z X
ZX
Laboratory for Reproductive Biology, University of North Carolina, Chapel Hill 27599.
Sar
M
M
Simental
J A
JA
Lane
M V
MV
Wilson
E M
EM
D000595:Amino Acid Sequence; D000728:Androgens / Q000378:metabolism; D000818:Animals; D001692:Biological Transport; D002467:Cell Nucleus / Q000378:metabolism*; D002478:Cells, Cultured; D004268:DNA-Binding Proteins / Q000378:metabolism*; D000882:Haplorhini; D006801:Humans; D008024:Ligands; D008969:Molecular Sequence Data; D016296:Mutagenesis; D021382:Protein Sorting Signals / Q000737:chemistry / Q000378:metabolism*; D011944:Receptors, Androgen / Q000737:chemistry / Q000378:metabolism*
D000728:Androgens; D004268:DNA-Binding Proteins; D008024:Ligands; D021382:Protein Sorting Signals; D011944:Receptors, Androgen