Abstract

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 NLS 383-386 and NLS 456-460 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.

Original languageEnglish (US)
Pages (from-to)532-541
Number of pages10
JournalExperimental Biology and Medicine
Volume234
Issue number5
DOIs
StatePublished - May 1 2009

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Nuclear Localization Signals
Cell proliferation
Cell Proliferation
Mutation
Bromodeoxyuridine
methionine-enkephalin receptor
Nuclear Pore
Nuclear Envelope
Protein Sorting Signals
Labeling
Cytoplasm
Cells

All Science Journal Classification (ASJC) codes

  • Biochemistry, Genetics and Molecular Biology(all)

Cite this

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title = "Dependence on nuclear localization signals of the opioid growth factor receptor in the regulation of cell proliferation",
abstract = "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 NLS 383-386 and NLS 456-460 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.",
author = "Fan Cheng and Patricia McLaughlin and Michael Verderame and Ian Zagon",
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AB - 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 NLS 383-386 and NLS 456-460 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.

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