Stimulation of Slow Skeletal Muscle Fiber Gene Expression by Calcineurinin Vivo

Francisco J. Naya, Brian Mercer, John Shelton, James A. Richardson, R. Sanders Williams, Eric N. Olson
2000 Journal of Biological Chemistry  
Adult skeletal muscle fibers can be categorized into fast and slow twitch subtypes based on specialized contractile and metabolic properties and on distinctive patterns of muscle gene expression. Muscle fiber-type characteristics are dependent on the frequency of motor nerve stimulation and are thought to be controlled by calcium-dependent signaling. The calcium, calmodulindependent protein phosphatase, calcineurin, stimulates slow fiber-specific gene promoters in cultured skeletal muscle
more » ... and the calcineurin inhibitor, cyclosporin A, inhibits slow fiber gene expression in vivo, suggesting a key role of calcineurin in activation of the slow muscle fiber phenotype. Calcineurin has also been shown to induce hypertrophy of cardiac muscle and to mediate the hypertrophic effects of insulin-like growth factor-1 on skeletal myocytes in vitro. To determine whether activated calcineurin was sufficient to induce slow fiber gene expression and hypertrophy in adult skeletal muscle in vivo, we created transgenic mice that expressed activated calcineurin under control of the muscle creatine kinase enhancer. These mice exhibited an increase in slow muscle fibers, but no evidence for skeletal muscle hypertrophy. These results demonstrate that calcineurin activation is sufficient to induce the slow fiber gene regulatory program in vivo and suggest that additional signals are required for skeletal muscle hypertrophy. Adult skeletal muscle fibers can be generally classified as fast or slow on the basis of their contractile and metabolic properties (1, 2). Certain fast fibers exhibit a glycolytic metabolism, whereas slow twitch fibers are predominantly oxidative. These properties reflect the expression of specific sets of fast and slow contractile protein isoforms of myosin heavy and light chains, tropomyosin, and troponins, as well as myoglobin. The fiber-type characteristics of adult skeletal muscle are regulated, at least in part, by the frequency of motor nerve stimulation and resulting changes in intracellular calcium lev-els (3-5). Tonic low frequency stimulation, either from the motor neuron or by direct electrical stimulation of muscle fibers, induces the slow fiber phenotype, whereas denervation or lack of electrical stimulation evokes a slow-to-fast fiber conversion (reviewed in Ref. 6). It is well established that slow fibers maintain higher levels of intracellular calcium (7, 8), but the signaling systems responsible for activation of the slow fiber gene program remain poorly understood. Calcineurin is a calcium-, calmodulin-dependent protein phosphatase activated by changes in intracellular calcium (reviewed in Ref. 9). The properties of calcineurin have been studied most extensively in T cells, whereupon activation in response to T cell receptor signaling, calcineurin dephosphorylates nuclear factor of activated T cells (NFAT) 1 proteins, which translocate to the nucleus and act combinatorially with other transcription factors (reviewed in Ref. 10). Calcineurin-dependent activation of NFAT is driven selectively by sustained, low amplitude elevations in intracellular calcium concentrations, while brief calcium transients, even of high amplitude, are insufficient for this purpose (11). Activation of calcineurin-dependent genes in T cells requires costimulation of mitogenactivated and calmodulin-dependent protein kinase signaling pathways (10), which activate the AP-1 and MEF2 transcription factors. Recent studies have also shown that activated calcineurin stimulates transcriptional activity of MEF2 transcription factors (12), expressed in T cells and muscle (reviewed in Ref. 13), through mechanisms that remain to be defined. Several properties of calcineurin signaling suggest that it may participate in the control of slow skeletal muscle gene expression. First, its selective activation by prolonged elevation of calcium and insensitivity to high amplitude calcium spikes correspond to the type of signaling thought to activate slow fiber-specific genes (9). Second, the control regions of several slow fiber-specific genes contain adjacent binding sites for . Third, transplant patients maintained on the calcineurin inhibitors, cyclosporin A (CsA) and FK-506, develop skeletal myopathy and a loss of skeletal muscle oxidative capacity, suggesting an important role for calcineurin in regulating muscle function (18) . Recently, we reported that calcineurin up-regulates slow muscle fiber genes in cultured C2C12 muscle cells (15). Transcriptional activation of the myoglobin and slow troponin I (TnI) genes by calcineurin was dependent on adjacent binding sites for NFAT and MEF2. Moreover, treatment of rats with CsA or FK-506 resulted in partial conversion of slow to fast fibers, suggesting that calcineurin activity was required for slow fiber gene expression. However, since systemic administration of CsA can also inhibit calcineurin activity in neurons, potentially altering motor innervation, and also has nonspecific effects, it was unclear whether the decrease in slow fiber gene expression in response to CsA reflected an essential role of calcineurin in skeletal muscle per se. Calcineurin has also been implicated in hypertrophy of cardiac and skeletal muscle (19 -21). In primary cardiomyocytes, calcineurin activation is required for hypertrophic growth in
doi:10.1074/jbc.275.7.4545 pmid:10671477 fatcat:l6bspdh7qvfcbfzibpszssii44