Analysis of muscle creatine kinase gene regulatory elements in skeletal and cardiac muscles of transgenic mice

DB Donoviel, MA Shield, JN Buskin… - … and cellular biology, 1996 - Am Soc Microbiol
DB Donoviel, MA Shield, JN Buskin, HS Haugen, CH Clegg, SD Hauschka
Molecular and cellular biology, 1996Am Soc Microbiol
Regulatory regions of the mouse muscle creatine kinase (MCK) gene, previously discovered
by analysis in cultured muscle cells, were analyzed in transgenic mice. The 206-bp MCK
enhancer at nt–1256 was required for high-level expression of MCK-chloramphenicol
acetyltransferase fusion genes in skeletal and cardiac muscle; however, unlike its behavior
in cell culture, inclusion of the 1-kb region of DNA between the enhancer and the basal
promoter produced a 100-fold increase in skeletal muscle activity. Analysis of enhancer …
Abstract
Regulatory regions of the mouse muscle creatine kinase (MCK) gene, previously discovered by analysis in cultured muscle cells, were analyzed in transgenic mice. The 206-bp MCK enhancer at nt–1256 was required for high-level expression of MCK-chloramphenicol acetyltransferase fusion genes in skeletal and cardiac muscle; however, unlike its behavior in cell culture, inclusion of the 1-kb region of DNA between the enhancer and the basal promoter produced a 100-fold increase in skeletal muscle activity. Analysis of enhancer control elements also indicated major differences between their properties in transgenic muscles and in cultured muscle cells. Transgenes in which the enhancer right E box or CArG element were mutated exhibited expression levels that were indistinguishable from the wild-type transgene. Mutation of three conserved E boxes in the MCK 1,256-bp 5′ region also had no effect on transgene expression in thigh skeletal muscle expression. All of these mutations significantly reduced activity in cultured skeletal myocytes. However, the enhancer AT-rich element at nt–1195 was critical for expression in transgenic skeletal muscle. Mutation of this site reduced skeletal muscle expression to the same level as transgenes lacking the 206-bp enhancer, although mutation of the AT-rich site did not affect cardiac muscle expression. These results demonstrate clear differences between the activity of MCK regulatory regions in cultured muscle cells and in whole adult transgenic muscle. This suggests that there are alternative mechanisms of regulating the MCK gene in skeletal and cardiac muscle under different physiological states.
American Society for Microbiology