Regulated Binding of Importin-α to Protein Kinase Cδ in Response to Apoptotic Signals Facilitates Nuclear Import

• Published in: The Journal of biological chemistry 2011-10, Vol.286 (41), p.35716-35724
• Main Authors: Adwan, Tariq S., Ohm, Angela M., Jones, David N.M., Humphries, Michael J., Reyland, Mary E.
• Format: Article
• Language: English
• Subjects: Active Transport, Cell Nucleus - physiology; alpha Karyopherins - genetics; alpha Karyopherins - metabolism; Amino Acid Motifs; Animals; Apoptosis; Apoptosis - physiology; Cell Line; Cell Nucleus - enzymology; Cell Nucleus - genetics; Cytoplasm - enzymology; Cytoplasm - genetics; Humans; Mice; Mutagenesis; Nuclear Localization Signals - genetics; Nuclear Localization Signals - metabolism; Nuclear Translocation; Phosphorylation - physiology; Phosphotyrosine; Protein Binding - physiology; Protein Kinase C (PKC); Protein Kinase C-delta - genetics; Protein Kinase C-delta - metabolism; Protein Phosphorylation; Rats; Signal Transduction
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M111.255950

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. Background: Tyrosine phosphorylation regulates nuclear translocation of proapoptotic protein kinase C delta (PKCδ). Results: Tyrosine phosphorylation causes a conformational change that exposes the nuclear localization sequence, allowing binding of importin-α. Conclusion: Nuclear localization of PKCδ is regulated by access of importin-α to the nuclear localization sequence. Significance: Nuclear import of PKCδ, which induces apoptosis, is tightly regulated so as to prevent inappropriate cell death.