Acetylation‐phosphorylation cross‐talk: A role for HDACs in the regulation of PKCdelta/theta phosphorylation

Abstract

Post‐translational modifications (PTMs) such as acetylation and phosphorylation have been implicated in contractile dysfunction and muscle wasting. Moreover, mounting evidence demonstrates that histone deacetylase (HDAC) inhibitors attenuate muscle wasting in animal models of disuse and muscular dystrophy. Historically, lysine acetylation was studied in the regulation of gene transcription, where for instance HDACs remove acetyl groups on lysine residues of histone proteins to suppress gene expression. However, recent evidence has shown that non‐histone proteins are also regulated by acetylation; functioning in protein stability, intracellular signaling, mitochondrial function and muscle contraction. Despite our current understanding for lysine acetylation and protein phosphorylation in muscle, how these PTMs cross‐talk to regulate muscle physiology remains unclear. In our attempt to understand acetylation‐phosphorylation cross‐talk, we examined the role for HDAC inhibitors on protein kinase C delta (PKCδ) and delta/theta (PKCδ/θ) phosphorylation. PKC is composed of multiple isoforms and its activation has been linked to muscle disease and dysfunction. Myotubes were treated with or without a pan‐HDAC inhibitor (trichostatin A; TSA) in the absence or presence of a pathological agonist (i.e. PMA). Phorbol 12‐myristate 13‐acetate (PMA) increased phosphorylation of PKCδ and PKCδ/θ. Interestingly, PMA‐induced PKCδ and PKCδ/θ phosphorylation was amplified in myotubes treated with the pan‐HDAC inhibitor TSA. Class selective HDAC inhibitors confirmed these findings and further demonstrated that inhibition of class I HDACs (HDAC1, 2, 3) was sufficient to increase PKCδ and PKCδ/θ phosphorylation. It should be noted that total PKCδ and PKCδ/θ protein expression increased with class I HDAC inhibition concomitant to the increase in protein phosphorylation. These data suggest that HDAC inhibition increases total PKC gene/protein expression that can then be phosphorylated. Finally, we report that the HDAC 1/2 selective inhibitor Romidepsin also increased total and phosphorylated PKCδ and PKCδ/θ in response to PMA. Genetic knockdown studies are currently underway to elucidate the specific HDAC that regulates PKC gene/protein expression, while chromatin immunoprecipitation (ChIP) is being used to determine HDAC binding to the PKC promoter. Lastly, elucidating a functional consequence for HDAC inhibitor‐mediated regulation of PKCδ and PKCδ/θ protein expression and phosphorylation in muscle could yield significant insight into muscle wasting and function.Support or Funding InformationThis work is supported by the USDA NIFA (Hatch‐NEV00727, Hatch‐NEV00767), the Dennis Meiss & Janet Ralston Fund for Nutri‐epigenetic Research, the National Institute for General Medical Sciences (NIGMS) of the NIH (P20 GM130459) and the National Heart, Lung, and Blood Institute of the NIH (R15 HL143496) and NSF EPSCOR Track II (OIA‐1826801) to B.S.F.