Skeletal muscle is composed of both slow-twich oxidative myofibers and fast-twitch glycolytic myofibers that differentially impact muscle metabolism, function, and eventually whole-body physiology. Here we show that the mesodermal transcription factor T-box 15 (Tbx15) is highly and specifically expressed in glycolytic myofibers. Ablation of Tbx15 in vivo leads to a decrease in muscle size due to a decrease in the number of glycolytic fibers, associated with a small increase in the number of oxidative fibers. This shift in fiber composition results in muscles with slower myofiber contraction and relaxation, and also decreases whole-body oxygen consumption, reduces spontaneous activity, increases adiposity, and glucose intolerance. Mechanistically, ablation of Tbx15 leads to activation of AMPK signaling and a decrease in Igf2 expression. Thus, Tbx15 is one of a limited number of transcription factors to be identified with a critical role in regulating glycolytic fiber identity and muscle metabolism.

A primary muscle cell culture derived from newborn rabbit muscle and growing on microcarriers in suspension was established. When cultured for several weeks, the myotubes in this model develop the completely adult pattern of fast myosin light and heavy chains. When Ca 2؉ ionophore is added to the culture medium on day 11, raising intracellular [Ca 2؉ ] about 10-fold, the myotubes develop to exhibit properties of an adult slow muscle by day 30, expressing slow myosin light as well as heavy chains, elevated citrate synthase, and reduced lactate dehydrogenase. The remarkable plasticity of these myotubes becomes apparent, when 8 days after withdrawal of the ionophore a marked slow-to-fast transition, as judged from the expression of isomyosins and metabolic enzymes, occurs.While the alterations that occur in mammalian muscle during fast-to-slow transition in vivo are known in great detail, mostly from chronic electrostimulation experiments with fast hindlimb muscles (1), the information on the mechanism initiating this process is sparse. A reduced intracellular phosphorylation potential and an elevated intracellular Ca 2ϩ concentration ([Ca 2ϩ ] i ), as they occur during sustained contractile activity, have been discussed as possible trigger events (2-5). It was our aim to test whether an increase in [Ca 2ϩ ] i imposed on the muscle cell is indeed able to induce a fast-to-slow transition. Because such an experiment cannot be performed in vivo, we were searching for a suitable myogenic cell culture system in which this would become feasible.Thus, this paper has two goals: (i) to report how a primary muscle culture can be grown that develops into an adult-like state, expressing adult myosin of the fast type only, and (ii) to study whether manipulation of [Ca 2ϩ ] i induces a shift between ''fast'' and ''slow'' fiber properties in the cultured myotubes. We show, first, that myotubes derived from newborn rabbit muscles and grown for 4 weeks on microcarriers in suspension possess a purely adult fast myosin pattern. Second, we show that these myotubes exhibit a similarly remarkable plasticity as it is known for adult rabbit muscles (1). This plasticity becomes apparent during exposure of the myotubes to high [Ca 2ϩ ] i , which causes development of the slow isoforms of myosin light chains (MLCs) and myosin heavy chains (MHCs) instead of their fast counterparts, of an increased aerobic metabolic capacity accompanied by a decreased anaerobic capacity as evidenced from elevated citrate synthase (CS) and reduced lactate dehydrogenase (LDH) levels, and of an increased activity ratio of the carbonic anhydrases CA III/CA II. Lowering [Ca 2ϩ ] i back to normal levels is followed by a reversal of all these changes within a few days. EXPERIMENTAL PROCEDURESCulture and Harvesting of Muscle Cells. Newborn White New Zealand rabbits were killed by decapitation. Hindlimb muscles were cut in small pieces and incubated in BSS, pH 7.0 (4.56 mM KCl͞0.44 mM KH 2 PO 4 ͞0.42 mM Na 2 HPO 4 ͞25 mM NaHCO 3 ͞ 119.8 mM NaCl͞50 mg/...

Carbonic anhydrase III is a cytosolic protein which is particularly abundant in skeletal muscle, adipocytes, and liver. The specific activity of this isozyme is quite low, suggesting that its physiological function is not that of hydrating carbon dioxide. To understand the cellular roles of carbonic anhydrase III, we inactivated the Car3 gene. Mice lacking carbonic anhydrase III were viable and fertile and had normal life spans. Carbonic anhydrase III has also been implicated in the response to oxidative stress. We found that mice lacking the protein had the same response to a hyperoxic challenge as did their wild-type siblings. No anatomic alterations were noted in the mice lacking carbonic anhydrase III. They had normal amounts and distribution of fat, despite the fact that carbonic anhydrase III constitutes about 30% of the soluble protein in adipocytes. We conclude that carbonic anhydrase III is dispensable for mice living under standard laboratory husbandry conditions.