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MUSCLE CELL BIOLOGY AND CELL MOTILITY

Real-time imaging of peroxisome proliferator-activated receptor- coactivator-1 promoter activity in skeletal muscles of living mice

Departments of ¹Medicine and ²Radiation Oncology, Duke University Medical Center, Durham, North Carolina 27710; and ³Department of Life Sciences, Graduate School of Arts and Sciences, The University of Tokyo, Komaba, Tokyo 305-8902, Japan

Submitted 2 October 2003 ; accepted in final form 13 May 2004

In response to sustained increase in contractile activity, mammalian skeletal muscle undergoes adaptation with enhanced mitochondrial biogenesis and fiber type switching. The peroxisome proliferator-activated receptor- coactivator-1 (PGC-1) was recently identified as a key regulator for these adaptive processes. To investigate the sequence elements in the PGC-1 gene that are responsible for activity-dependent transcriptional activation, we have established a unique system to analyze promoter activity in skeletal muscle of living mice. Expression of PGC-1-firefly luciferase reporter gene in mouse tibialis anterior muscle transfected by electric pulse-mediated gene transfer was assessed repeatedly in the same muscle by using optical bioluminescence imaging analysis before and after low-frequency (10 Hz) motor nerve stimulation. Nerve stimulation (2 h) resulted in a transient 3-fold increase (P < 0.05) in PGC-1 promoter activity along with a 1.6-fold increase (P < 0.05) in endogenous PGC-1 mRNA. Mutation of two consensus myocyte enhancer factor 2 (MEF2) binding sites (–2901 and –1539) or a cAMP response element (CRE) (–222) completely abolished nerve stimulation-induced increase in PGC-1 promoter activity. These findings provide direct evidence that contractile activity-induced PGC-1 promoter activity in skeletal muscle is dependent on the MEF2 and the CRE sequence elements. The experimental methods used in the present study have general applicability to studies of gene regulation in muscle.

adaptation; fiber type switching; mitochondrial biogenesis; electric pulse-mediated gene transfer; optical bioluminescence imaging

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