Opposite Effect of Intracellular Ca2+ and Protein Kinase C on the Expression of Inwardly Rectifying K+Channel 1 in Mouse Skeletal Muscle

Abstract

The level of inwardly rectifying K ؉ channel 1 (IRK1) mRNA decreased upon denervation and increased during muscle differentiation in mouse skeletal muscle. To identify the mechanism(s) underlying the regulation of IRK1 mRNA expression, we examined its expression using the well differentiated C2C12 mouse skeletal muscle cell line as a model system. Since nerve-induced muscle activity results in contraction, it was questioned whether the changes in IRK1 expression might be relevant to the increased intracellular calcium that functions as a cytoplasmic messenger in excitation-contraction coupling. Indeed, activation of either L-type calcium channels or ryanodine receptors increased the level of IRK1 mRNA. More directly, ionomycin activated the IRK1 expression in time-and dose-dependent manners, which was abolished by treatment with EGTA. Genistein, a tyrosine kinase inhibitor, also abolished the stimulating effect of ionomycin. Meanwhile, activation of protein kinase C by 12-O-tetradecanoylphorbol acetate (TPA) markedly decreased the level of IRK1 mRNA, which required ongoing protein synthesis. Actinomycin D experiments revealed that ionomycin increased the half-life of IRK1 mRNA from 0.86 to 1.97 h, but TPA decreased it to 0.38 h. However, neither ionomycin nor TPA appreciably altered the rate of IRK1 gene transcription. Based on these observations, we conclude that intracellular calcium and protein kinase C are oppositely involved in the muscle activity-dependent regulation of IRK1 gene expression and that both act at the level of mRNA stability.It has been known that denervation influences many biophysical and biochemical properties of skeletal muscle fibers (1-7). The mechanisms by which denervation initiates these changes are still unclear. They may be caused by the loss of neurotrophic factors normally released from the nerve terminals (8, 9). Alternatively, the electrical inactivity of the denervated muscle might be responsible, since direct electrical stimulation to the denervated muscle restores all passive electrical parameters of the membrane that were observed without denervation (10, 11). Considering the importance of calcium in the process of muscle contraction induced by neural activity, calcium may play a critical role in linking the biochemical and biophysical changes with muscle activity.Calcium is known to be involved in many cellular events as a second messenger. A regulatory role of calcium in the expression of sodium channels, acetylcholinesterase, and nicotinic acetylcholine receptor has been suggested (12-16). In addition, it has recently been shown that calcium influx blocks the expression of nicotinic acetylcholine receptor ␣-subunit gene in chick skeletal muscle (17). There are reports that coupling of the electrical activity with altered gene expression is mediated by protein kinase C (PKC) 1 pathway (18, 19). Gonoi and Hasegawa (20) demonstrate by using a patch clamp method that innervation of skeletal muscle fibers plays a key role in the induction and maintenance of inwardly rectifyin...

Reduction in Intracellular Calcium Levels Inhibits Myoblast Differentiation

“…However, the intracellular pathways that transduce these signals to the nucleus are only beginning to be understood. Previous reports have suggested that calcium-activated signaling cascades may play an important role in intracellular signaling during myocyte differentiation (7,11,13,25,26). Using C2C12 cells as a model of myocyte differentiation, we demonstrate that the maintenance of normal intracellular calcium concentrations is vital to myocyte gene expression and differentiation.…”

“…For example, calpain-mediated protein degradation appears to allow cell fusion and differentiation of C2C12 cells (13). In addition, increased levels of intracellular calcium appear to stabilize mRNA encoding the inward-rectifying potassium channel 1 (26).…”

Ion Channel Development, Spontaneous Activity, and Activity-Dependent Development in Nerve and Muscle Cells

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Suppression of Kir2.3 Activity by Protein Kinase C Phosphorylation of the Channel Protein at Threonine 53

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