PKCα:: a versatile key for decoding the cellular calcium toolkit

Conventional protein kinases C (cPKCs) play an essential role in signal transduction and are believed to integrate both global Ca2+ transients and diacylglycerol signals. We provide evidence that PKC alpha is a ubiquitous readout sensor for the cellular Ca2+ toolkit, including highly restricted elementary Ca2+ release. Threshold stimulations of cells with Ca2+-mobilizing agonists resulted in PKCa translocation events with limited spatial spreads (< 4 mu m) comprising two groups of lifetimes; brief events (400-1,500 ms) exclusively mediated by Ca2+-C2 domain membrane interactions and long-lasting events (> 4 s) resulting from longer DAG-C1a domain-mediated membrane interactions. Although upon uncaging NP-EGTA, which is a caged Ca2+ compound, WT-PKC alpha displayed rapid membrane translocations within < 250 ms, PKCa constructs with C2 domains mutated in their Ca2+-binding region lacked any Ca2+-dependent translocation. Flash photolysis of diazo-2, a photosensitive caged Ca2+ buffer, revealed a biphasic membrane dissociation ( slow and fast period) of WT-PKC alpha. The slow phase was absent in cells expressing PKC alpha-constructs containing mutated C1a-domains with largely reduced DAG binding. Thus, two groups of PKCa membrane interactions coexist; C2- and C1a-mediated interactions with different lifetimes but rapid interconversion. We conclude that PKCa can readout very fast and, spatially and temporally, very complex cellular Ca2+ signals. Therefore, cPKCs are important transducers for the ubiquitous cellular Ca2+ signaling toolkit.