Calcium-regulated changes in mitochondrial phenotype in skeletal muscle cells

Cytochrome c expression and mitochondrial biogenesis can be invoked by elevated intracellular Ca2+ in muscle cells. To characterize the potential role of Ca2+ as a messenger involved in mitochondrial biogenesis in muscle, we determined the effects of the Ca2+ ionophore A-23187 on the expression of nuclear- and mitochondrially encoded genes. Treatment of myotubes with 1 muM A-23187 for 48-96 h increased nuclear-encoded beta-subunit F(1)ATPase and malate dehydrogenase (MDH) mRNA levels by 50-100% ( P < 0.05) but decreased mRNA levels of glutamate dehydrogenase (GDH) by 19% (P < 0.05). mRNA levels of the cytochrome c oxidase (COX) nuclear-encoded subunits IV, Vb, and VIc were unchanged, whereas the mitochondrially encoded subunits COX II and COX III were decreased by 30 and 70%, respectively ( P < 0.05). This was paralleled by a 20% decrease ( P < 0.05) in COX activity. These data suggest that cytoplasmic Ca2+ differentially regulates the mRNA level of nuclear and mitochondrial genes. The decline in COX II and III mRNA may be mediated by Tfam, because A-23187 modestly reduced Tfam levels by 48 h. A-23187 induced time-dependent increases in Egr-1 mRNA, along with the activation of ERK1/2 and AMP-activated protein kinase. MEK inhibition with PD-98059 attenuated the increase in Egr-1 mRNA. A-23187 also increased Egr-1, serum response factor, and Sp1 protein expression, transcription factors implicated in mitochondrial biogenesis. Egr-1 overexpression increased nuclear- encoded cytochrome c transcriptional activation by 1.5-fold (P < 0.05) and reduced GDH mRNA by 37% ( P < 0.05) but had no effect on MDH or beta-subunit F1ATPase mRNA. These results indicate that changes in intracellular Ca2+ can modify mitochondrial phenotype, in part via the involvement of Egr-1.