DNA binding domains in diverse nuclear receptors function as nuclear export signals.
BACKGROUND
The nuclear receptor superfamily of transcription factors directs gene expression through DNA sequence-specific interactions with target genes. Nuclear import of these receptors involves recognition of a nuclear localization signal (NLS) by importins, which mediate translocation into the nucleus. Nuclear receptors lack a leucine-rich nuclear export signal (NES), and export is insensitive to leptomycin B, indicating that nuclear export is not mediated by Crm1.


RESULTS
We set out to define the NES in the glucocorticoid receptor (GR) and to characterize the export pathway. We found that the 69 amino acid DNA binding domain (DBD) of GR, which is unrelated to any known NES, is necessary and sufficient for export. Mutational analysis revealed that a 15 amino acid sequence between the two zinc binding loops in the GR-DBD confers nuclear export to a GFP reporter protein, and alanine-scanning mutagenesis was used to identify the residues within this sequence that are critical for export. The DBD is highly related (41%-88% identity) in steroid, nonsteroid, and orphan nuclear receptors, and we found that the DBDs from ten different nuclear receptors all function as export signals. DBD-dependent nuclear export is saturable, and prolonged nuclear localization of the GR increases its transcriptional activity.


CONCLUSIONS
Multiple members of the nuclear receptor superfamily use a common pathway to exit the nucleus. We propose that NLS-mediated import and DBD-mediated export define a shuttling cycle that integrates the compartmentalization and activity of nuclear receptors.
Black
B E
BE
Center for Cell Signaling, University of Virginia, Charlottesville, VA 22908, USA.
Holaska
J M
JM
Rastinejad
F
F
Paschal
B M
BM
D021581:Active Transport, Cell Nucleus; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D019556:COS Cells; D002135:Calcium-Binding Proteins / Q000378:metabolism; D037282:Calreticulin; D002467:Cell Nucleus / Q000378:metabolism*; D002522:Chlorocebus aethiops; D004247:DNA / Q000378:metabolism*; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D019913:Nuclear Localization Signals; D011965:Receptors, Glucocorticoid / Q000235:genetics / Q000378:metabolism*; D012261:Ribonucleoproteins / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D015398:Signal Transduction; D014158:Transcription, Genetic; D016335:Zinc Fingers
D002135:Calcium-Binding Proteins; D037282:Calreticulin; D019913:Nuclear Localization Signals; D011965:Receptors, Glucocorticoid; D012261:Ribonucleoproteins; D004247:DNA