Reduced protein kinase C delta association with a higher molecular weight complex in mitochondria of Barth Syndrome lymphoblasts

Protein kinase C delta (PKCδ) is a signaling kinase that regulates several cellular responses and is controlled via multi-site phosphorylation. The PKCδ signalosome exists as a high molecular weight complex in mitochondria and adjusts the fuel flux from glycolytic sources to the intensity of mitochondrial respiration, thus controlling mitochondrial oxidative phosphorylation. In the X-linked genetic disease Barth Syndrome (BTHS) mitochondrial oxidative phosphorylation is impaired due to a mutation in the TAFAZZIN gene which results in reduction in the phospholipid cardiolipin (CL) and an accumulation of monolysocardiolipin (MLCL). We previously demonstrated, through phosphokinome analysis, that phosphorylation of PKCδ is altered on multiple sites in BTHS patient lymphoblasts. In this study, we examined if PKCδ association with a higher molecular weight complex was altered in mitochondria of BTHS lymphoblasts. BTHS lymphoblasts exhibited reduction in all molecular species of CL examined and accumulation of trioleoyl-MLCL. Immunoblot analysis of blue native-polyacrylamide gel electrophoresis mitochondrial fractions revealed that PKCδ was associated with a higher molecular weight complex and that this was markedly reduced in BTHS patient B lymphoblasts compared to controls in spite of an increase in PKCδ protein expression. The elevated expression level of PKCδ in BTHS lymphoblasts was associated with increased citrate synthase activity indicative of abnormal mitochondrial proliferation. We hypothesize that the lack of PKCδ within this higher molecular weight complex may contribute to defective mitochondrial PKCδ signaling and thus to the bioenergetic defects observed in BTHS.

The mono-ADP-ribosyltransferase ARTD10 regulates the voltage-gated K+ channel Kv1.1 through protein kinase C delta

Background ADP-ribosylation is a ubiquitous post-translational modification that involves both mono- and poly-ADP-ribosylation. ARTD10, also known as PARP10, mediates mono-ADP-ribosylation (MARylation) of substrate proteins. A previous screen identified protein kinase C delta (PKCδ) as a potential ARTD10 substrate, among several other kinases. The voltage-gated K+ channel Kv1.1 constitutes one of the dominant Kv channels in neurons of the central nervous system and the inactivation properties of Kv1.1 are modulated by PKC. In this study, we addressed the role of ARTD10-PKCδ as a regulator of Kv1.1. Results We found that ARTD10 inhibited PKCδ, which increased Kv1.1 current amplitude and the proportion of the inactivating current component in HeLa cells, indicating that ARTD10 regulates Kv1.1 in living cells. An inhibitor of ARTD10, OUL35, significantly decreased peak amplitude together with the proportion of the inactivating current component of Kv1.1-containing channels in primary hippocampal neurons, demonstrating that the ARTD10-PKCδ signaling cascade regulates native Kv1.1. Moreover, we show that the pharmacological blockade of ARTD10 increases excitability of hippocampal neurons. Conclusions Our results, for the first time, suggest that MARylation by ARTD10 controls neuronal excitability.