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

PKC delta translocates into the nucleus in response to apoptotic agents and functions as a potent cell death signal. Cytoplasmic retention of PKC delta and its transport into the nucleus are essential for cell homeostasis, but how these processes are regulated is poorly understood. We show that PKC delta resides in the cytoplasm in a conformation that precludes binding of importin-alpha. A structural model of PKC delta in the inactive state suggests that the nuclear localization sequence (NLS) is prevented from binding to importin-alpha through intramolecular contacts between the C2 and catalytic domains. We have previously shown that PKC delta is phosphorylated on specific tyrosine residues in response to apoptotic agents. Here, we show that phosphorylation of PKC delta at Tyr-64 and Tyr-155 results in a conformational change that allows exposure of the NLS and binding of importin-alpha. In addition, Hsp90 binds to PKC delta with similar kinetics as importin-alpha and is required for the interaction of importin-alpha with the NLS. Finally, we elucidate a role for a conserved PPxxP motif, which overlaps the NLS, in nuclear exclusion of PKC delta. Mutagenesis of the conserved prolines to alanines enhanced importin-alpha binding to PKC delta and induced its nuclear import in resting cells. Thus, the PPxxP motif is important for maintaining a conformation that facilitates cytosplasmic retention of PKC delta. Taken together, this study establishes a novel mechanism that retains PKC delta in the cytoplasm of resting cells and regulates its nuclear import in response to apoptotic stimuli.