Dependence on nuclear localization signals of the opioid growth factor receptor in the regulation of cell proliferation.
The opioid growth factor receptor (OGFr) mediates the inhibitory action of OGF on cell replication of normal and neoplastic cells. The spatiotemporal course of OGFr nucleocytoplasmic trafficking was determined with a probe of full-length OGFr fused to enhanced green fluorescent protein (eGFP). Translation of OGFr required 8.5 hours, and transit into the nucleus required 8 hours; OGFr remained in the nucleus for 8 days. OGFr was initially expressed on the outer nuclear envelope, transited to the paranuclear cytoplasm, and into the nucleus. Transport through the nuclear pore was elucidated by mutation of the nuclear localization signal (NLS) sequences in full-length OGFr. Mutation of each NLS reduced nuclear localization by 5%-50%, whereas simultaneous mutation of NLS383-386 and NLS456-460 abolished OGFr-eGFP nuclear localization in 80% of the cells. To determine whether intact NLSs are important for the inhibition of cell proliferation, DNA synthesis was monitored with BrdU. Wild-type OGFr-eGFP-transfected cells had 20% BrdU-positive cells, whereas cells with simultaneous mutation of all three NLS sites had a 70% labeling index. These results indicate that the regulation of cell proliferation by the OGF-OGFr axis is dependent on nucleocytoplasmic translocation and reliant on the integrity of two NLSs in OGFr to interact with transport receptors.
Cheng
Fan
F
Department of Neural and Behavioral Sciences, The Pennsylvania State University College of Medicine, H109, The Milton S. Hershey Medical Center, 500 University Drive, Room C3729, Hershey, PA 17033, USA.
McLaughlin
Patricia J
PJ
Verderame
Michael F
MF
Zagon
Ian S
IS
D021581:Active Transport, Cell Nucleus / Q000502:physiology; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D004247:DNA / Q000096:biosynthesis* / Q000235:genetics; D006801:Humans; D009154:Mutation; D009685:Nuclear Envelope / Q000235:genetics / Q000378:metabolism*; D019913:Nuclear Localization Signals / Q000235:genetics / Q000378:metabolism*; D014176:Protein Biosynthesis / Q000502:physiology*; D011957:Receptors, Opioid / Q000096:biosynthesis* / Q000235:genetics; D013997:Time Factors
D019913:Nuclear Localization Signals; D011957:Receptors, Opioid; C033593:methionine-enkephalin receptor; D004247:DNA