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School of Kinesiology, University of Illinois, Chicago, Illinois 60608
It has been well
established that expression of slow contractile protein genes in
skeletal muscle is regulated, in part, by activity from slow
motoneurons. However, very little is understood about the mechanism by
which neural activity regulates transcription of slow isoform genes.
The purpose of this investigation was first to more fully define the in
vivo DNA injection technique for use in both fast-twitch and
slow-twitch muscles and second to use the injection
technique for the identification of slow nerve-dependent regions of the
myosin light chain 2 slow (MLC2s)
gene. Initial experiments determined that the same amount of plasmid
DNA was taken up by both the slow-twitch soleus and fast-twitch
extensor digitorum longus (EDL) muscles and that injection of from 0.5 to 10 µg DNA/muscle is ideal for analysis of promoter activity during
regeneration. This technique was subsequently used to identify that the
region from 
800 to +12 base pairs of
MLC2s gene directed ~100 times
higher activity in the innervated soleus than in innervated EDL,
denervated soleus, or denervated EDL muscles. Placing the introns
upstream of either the MLC2s or SV40
promoter increased expression 5- and 2.7-fold, respectively, in
innervated soleus but not in innervated EDL, denervated soleus, or
denervated EDL muscles. These results demonstrate that
1) in vivo DNA injection is a
sensitive assay for promoter analysis in both fast-twitch and
slow-twitch skeletal muscles and 2)
both 5' flanking and intronic regions of the
MLC2s gene can independently and
synergistically direct slow nerve-dependent transcription in vivo.
in vivo deoxyribonucleic acid injection; muscle regeneration; contractile protein genes
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