Protein kinase C-δ regulates the subcellular localization of Shc in H2O2-treated cardiomyocytes

Guo J, Cong L, Rybin VO, Gertsberg Z, Steinberg SF. Protein kinase C-delta regulates the subcellular localization of Shc in H2O2-treated cardiomyocytes. Am J Physiol Cell Physiol 299: C770-C778, 2010. First published August 4, 2010; doi:10.1152/ajpcell.00170.2010.-Protein kinase C-delta (PKC delta) exerts important cardiac actions as a lipid-regulated kinase. There is limited evidence that PKC delta also might exert an additional kinase-independent action as a regulator of the subcellular compartmentalization of binding partners such as Shc (Src homologous and collagen), a family of adapter proteins that play key roles in growth regulation and oxidative stress responses. This study shows that native PKC delta forms complexes with endogenous Shc proteins in H2O2-treated cardiomyocytes; H2O2 treatment also leads to the accumulation of PKC delta and Shc in a detergent-insoluble cytoskeletal fraction and in mitochondria. H2O2-dependent recruitment of Shc isoforms to cytoskeletal and mitochondrial fractions is amplified by wild-type-PKC delta overexpression, consistent with the notion that PKC delta acts as a signal-regulated scaffold to anchor Shc in specific subcellular compartments. However, overexpression studies with kinase-dead (KD)PKC delta-K376R (an ATP-binding mutant of PKC delta that lacks catalytic activity) are less informative, since KD-PKC delta-K376R aberrantly localizes as a constitutively tyrosine-phosphorylated enzyme to detergent-insoluble and mitochondrial fractions of resting cardiomyocytes; relatively little KD-PKC delta-K376R remains in the cytosolic fraction. The aberrant localization and tyrosine phosphorylation patterns for KD-PKC delta-K376R do not phenocopy the properties of native PKC delta, even in cells chronically treated with GF109203X to inhibit PKC delta activity. Hence, while KD-PKC delta-K376R overexpression increases Shc localization to the detergent-insoluble and mitochondrial fractions, the significance of these results is uncertain. Our studies suggest that experiments using KD-PKC delta-K376R overexpression as a strategy to competitively inhibit the kinase-dependent actions of native PKC delta or to expose the kinase-independent scaffolding functions of PKC delta should be interpreted with caution.