PKCä signaling to mitochondria has been implicated in both mitochondrial apoptosis and metabolism. However, the mechanism by which PKCä interacts with mitochondria is not well understood. Using FRET-based imaging, we show that PKCä interacts with mitochondria by a novel and isozyme-specific mechanism distinct from its canonical recruitment to other membranes such as the plasma membrane or Golgi. Specifically, we show that PKCä interacts with mitochondria following stimulation with phorbol esters or, in L6 myocytes, with insulin via a mechanism that requires two steps. In the first step, PKCä translocates acutely to mitochondria by a mechanism that requires its C1A and C1B domains and a Leu-Asn sequence in its turn motif. In the second step, PKCä is retained at mitochondria by a mechanism that depends on its C2 domain, a unique Glu residue in its activation loop, intrinsic catalytic activity, and the mitochondrial membrane potential. In contrast, of these determinants, only the C1B domain is required for the phorbol ester-stimulated translocation of PKCä to other membranes. PKCä also basally localizes to mitochondria and increases mitochondrial respiration via many of the same determinants that promote its agonist-evoked interaction. PKCä localized to mitochondria has robust activity, as revealed by a FRET reporter of PKCä-specific activity (äCKAR). These data support a model in which multiple determinants unique to PKCä drive a specific interaction with mitochondria that promotes mitochondrial respiration.
Amino Acid Sequence Animals