Abstract
Peroxisome proliferator-activated receptor gamma co-activator 1alpha (PGC-1alpha) promotes mitochondrial biogenesis and slow fiber formation in skeletal muscle. We hypothesized that activation of the p38 mitogen-activated protein kinase (MAPK) pathway in response to increased muscle activity stimulated Pgc-1alpha gene transcription as part of the mechanisms for skeletal muscle adaptation. Here we report that a single bout of voluntary running induced a transient increase of Pgc-1alpha mRNA expression in mouse plantaris muscle, concurrent with an activation of the p38 MAPK pathway. Activation of the p38 MAPK pathway in cultured C2C12 myocytes stimulated Pgc-1alpha promoter activity, which could be blocked by the specific inhibitors of p38, SB203580 and SB202190, or a dominant negative p38. Furthermore, the p38-mediated increase in Pgc-1alpha promoter activity was enhanced by increased expression of the downstream transcription factor ATF2 and completely blocked by ATF2DeltaN, a dominant negative ATF2. Skeletal muscle-specific expression of a constitutively active activator of p38, MKK6E, in transgenic mice resulted in enhanced Pgc-1alpha and cytochrome oxidase IV protein expression in fast-twitch skeletal muscles. These findings suggest that contractile activity-induced activation of the p38 MAPK pathway promotes Pgc-1alpha gene expression and skeletal muscle adaptation.
Highlights
Adult skeletal muscle is remarkably plastic [1, 2]
These findings suggest that contractile activity-induced activation of the p38 mitogen-activated protein kinase (MAPK) pathway promotes Pgc-1␣ gene expression and skeletal muscle adaptation
It is currently unknown whether exercise-induced PGC-1␣ expression in skeletal muscle is essential for enhanced mitochondrial biogenesis and/or IIb-to-IIa fiber type switching, the latter being different from the reported function of PGC-1␣ in transgenic mice in promoting more type I fiber formation [6]
Summary
Animals—Adult (8-week-old) male C57BL/6J mice (The Jackson Laboratory) were housed in temperature-controlled (21 °C) quarters with a 12-h light/12-h dark cycle and provided with water and food (Purina Chow) ad libitum. Semiquantitative RT-PCR analysis was performed to measure endogenous PGC-1␣ mRNA expression in plantaris muscle in response to voluntary running as described previously [12, 35]. To detect FLAG-tagged MKK6E transgene expression in cultured C2C12 myocytes, ϳ15 g of proteins in total cell lysate for each sample was resolved by SDS-PAGE and probed with rabbit anti-FLAG antibody (F-7425, Sigma) with a dilution of 1:1000 as described above. The expected 1.5-kb PCR fragment was inserted into pTarget vector (Promega) to generate plasmid CMV-ATF2, in which ATF2 expression is under the control of the constitutively active CMV promoter. In the experiments with transfected C2C12 myocytes with MCK-MKK6E, myogenic differentiation was induced in C2C12 cells for 4 days by changing the culture medium to Dulbecco’s modified Eagle’s medium containing 2% horse serum. Statistical significance (p Ͻ 0.05) was determined by Student’s t test or analysis of variance followed by the Dunnett test for comparisons between two groups or multiple groups, respectively
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