Human Scythe contains a functional nuclear localization sequence and remains in the nucleus during staurosporine-induced apoptosis.
Human Scythe (also known as BAT3) has been implicated in the control of apoptosis and regulating heat shock protein (HSP) 70 activity. We have attempted to further characterize the role of human Scythe in HeLa cells by studying the cellular localization and functional domains of a hemagglutinin (HA) epitope-tagged Scythe protein. Several HA-Scythe deletion mutant proteins were expressed in HeLa cells and their localization was detected using indirect immunofluorescence. Our data demonstrate that full-length human Scythe is a nuclear protein that contains an active C-terminal nuclear localization sequence (NLS). Site-directed mutagenesis of the NLS leads to complete nuclear exclusion of full-length Scythe. Furthermore, induction of apoptosis by staurosporine does not cause redistribution or cleavage of Scythe, suggesting that Scythe remains localized in the nucleus during apoptosis. These results provide evidence that Scythe is a nuclear protein that probably does not interact with elements of the apoptotic machinery in the cytosol.
Manchen
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Department of Biological Sciences, University of Windsor, Windsor, Ontario, N9B 3P4, Canada.
Hubberstey
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AV
D000595:Amino Acid Sequence; D017209:Apoptosis; D002451:Cell Compartmentation; D019084:Fluorescent Antibody Technique, Indirect; D006367:HeLa Cells; D006801:Humans; D020287:In Situ Nick-End Labeling; D018832:Molecular Chaperones; D008969:Molecular Sequence Data; D019913:Nuclear Localization Signals / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D009687:Nuclear Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D011506:Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D019311:Staurosporine / Q000494:pharmacology
C092009:BAG6 protein, human; D018832:Molecular Chaperones; D019913:Nuclear Localization Signals; D009687:Nuclear Proteins; D011506:Proteins; D019311:Staurosporine