title: Regulated binding of importin-α to protein kinase Cδ in response to apoptotic signals facilitates nuclear import.
abstract: PKCδ translocates into the nucleus in response to apoptotic agents and functions as a potent cell death signal. Cytoplasmic retention of PKCδ and its transport into the nucleus are essential for cell homeostasis, but how these processes are regulated is poorly understood. We show that PKCδ resides in the cytoplasm in a conformation that precludes binding of importin-α. A structural model of PKCδ in the inactive state suggests that the nuclear localization sequence (NLS) is prevented from binding to importin-α through intramolecular contacts between the C2 and catalytic domains. We have previously shown that PKCδ is phosphorylated on specific tyrosine residues in response to apoptotic agents. Here, we show that phosphorylation of PKCδ at Tyr-64 and Tyr-155 results in a conformational change that allows exposure of the NLS and binding of importin-α. In addition, Hsp90 binds to PKCδ with similar kinetics as importin-α and is required for the interaction of importin-α with the NLS. Finally, we elucidate a role for a conserved PPxxP motif, which overlaps the NLS, in nuclear exclusion of PKCδ. Mutagenesis of the conserved prolines to alanines enhanced importin-α binding to PKCδ and induced its nuclear import in resting cells. Thus, the PPxxP motif is important for maintaining a conformation that facilitates cytosplasmic retention of PKCδ. Taken together, this study establishes a novel mechanism that retains PKCδ in the cytoplasm of resting cells and regulates its nuclear import in response to apoptotic stimuli.
authors.affiliation: Program in Cell Biology, Stem Cells and Development, University of Colorado, Anschutz Medical Campus, Aurora, Colorado 80045.
authors.forename: Tariq S
authors.initials: TS
authors.lastname: Adwan
authors.affiliation: Department of Craniofacial Biology, School of Dental Medicine, University of Colorado, Anschutz Medical Campus, Aurora, Colorado 80045.
authors.forename: Angela M
authors.initials: AM
authors.lastname: Ohm
authors.affiliation: Department of Pharmacology, School of Medicine, University of Colorado, Anschutz Medical Campus, Aurora, Colorado 80045.
authors.forename: David N M
authors.initials: DNM
authors.lastname: Jones
authors.affiliation: Program in Cell Biology, Stem Cells and Development, University of Colorado, Anschutz Medical Campus, Aurora, Colorado 80045.
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Humphries
authors.affiliation: Program in Cell Biology, Stem Cells and Development, University of Colorado, Anschutz Medical Campus, Aurora, Colorado 80045; Department of Craniofacial Biology, School of Dental Medicine, University of Colorado, Anschutz Medical Campus, Aurora, Colorado 80045. Electronic address: Mary.Reyland@UCDenver.edu.
authors.forename: Mary E
authors.initials: ME
authors.lastname: Reyland
mesh_terms: D021581:Active Transport, Cell Nucleus / Q000502:physiology; D020816:Amino Acid Motifs; D000818:Animals; D017209:Apoptosis / Q000502:physiology*; D002460:Cell Line; D002467:Cell Nucleus / Q000201:enzymology* / Q000235:genetics; D003593:Cytoplasm / Q000201:enzymology* / Q000235:genetics; D006801:Humans; D051379:Mice; D016296:Mutagenesis; D019913:Nuclear Localization Signals / Q000235:genetics / Q000378:metabolism; D010766:Phosphorylation / Q000502:physiology; D011485:Protein Binding / Q000502:physiology; D051745:Protein Kinase C-delta / Q000235:genetics / Q000378:metabolism*; D051381:Rats; D028901:alpha Karyopherins / Q000235:genetics / Q000378:metabolism*
chemical_list: D019913:Nuclear Localization Signals; D028901:alpha Karyopherins; D051745:Protein Kinase C-delta