Differences in the function of three conserved E-boxes of the muscle creatine kinase gene in cultured myocytes and in transgenic mouse skeletal and cardiac muscle.
The 1256-base pair enhancer-promoter of the mouse muscle creatine kinase gene includes three CAnnTG E-boxes that are conserved among mammals and have flanking and middle sequences conforming to consensus muscle regulatory factor binding sites. This study seeks to determine whether these E-boxes are critical for muscle creatine kinase expression in physiologically distinct muscles. Mutations of the "right" and "left" E-boxes in the enhancer decreased expression in cultured skeletal myocytes approximately 10- and 2-fold, respectively, whereas a "promoter" E-box mutation had little effect. In neonatal myocardiocytes, the left E-box mutation decreased expression approximately 3-fold, whereas right or promoter E-box mutations had no effect. Very different effects were seen in transgenic mice, where the promoter E-box mutation decreased expression in quadriceps, extensor digitorum longus, and soleus approximately 10-fold, and approximately 100-fold in distal tongue, diaphragm, and ventricle. The right E-box mutation, tested in the presence of the other two mutations, caused a significant decrease in distal tongue, but not in quadriceps, extensor digitorum longus, soleus, or ventricle. Mutation of the left E-box actually raised expression in soleus, suggesting a possible repressor role for this control element. The discrepancies between mutation effects in differentiating skeletal muscle cultures, neonatal myocardiocytes, and adult mice suggested that the E-boxes might play different roles during muscle development and adult steady-state function. However, transgenic analysis of embryonic and early postnatal mice indicated no positive role for these three E-boxes in early development, implying that differences in E-box function between adult muscle and cultured cells are the result of physiological signals.
Nguyen
Quynh-Giao V
QG
Department of Biochemistry, University of Washington, Seattle, WA 98195-7350, USA.
Buskin
Jean N
JN
Himeda
Charis L
CL
Shield
Margaret A
MA
Hauschka
Stephen D
SD
D000818:Animals; D000831:Animals, Newborn; D001483:Base Sequence; D002478:Cells, Cultured; D017124:Conserved Sequence; D003402:Creatine Kinase / Q000235:genetics*; D024721:E-Box Elements / Q000235:genetics / Q000502:physiology*; D004622:Embryo, Mammalian; D018507:Gene Expression Regulation, Developmental / Q000235:genetics; D015971:Gene Expression Regulation, Enzymologic / Q000235:genetics; D051379:Mice; D008822:Mice, Transgenic; D032342:Muscle Cells / Q000201:enzymology*; D018482:Muscle, Skeletal / Q000201:enzymology*; D009154:Mutation; D009206:Myocardium / Q000201:enzymology*; D051381:Rats; D017207:Rats, Sprague-Dawley
D003402:Creatine Kinase