Identification and characterization of a factor that binds to two human sarcomeric actin promoters.
A gel mobility shift assay was used to examine nuclear extracts of muscle and nonmuscle cell lines for a factor or factors that interact with the functional promoter segment of both the human cardiac and skeletal alpha-actin genes. A single major band of altered mobility was seen with nuclear extracts from C2 myoblasts and myotubes and L8 myoblasts and myotubes as well as several non-muscle cell lines. Competition experiments with nuclear extracts from both muscle and non-muscle cells localized the region of binding to a 10-base pair sequence of the cardiac alpha-actin gene containing the functional CArG box promoter domain. Similar results and binding site specificities were obtained with a DNA fragment from the human skeletal alpha-actin promoter. The CArG box binding factor was shown to be distinct from a CAAT box binding factor. A molecular weight of 67 kDa for the CArG box binding factor was determined by photoaffinity labeling of the complex and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Protease treatment resulted in a smaller fragment that still bound the cardiac alpha-actin promoter fragment and may represent a structurally distinct DNA-binding domain. We discuss how muscle-specific expression of the sarcomeric alpha-actin genes might be achieved with a ubiquitous transcription factor.
Boxer
L M
LM
Department of Medicine, Stanford University School of Medicine, Palo Alto, California.
Miwa
T
T
Gustafson
T A
TA
Kedes
L
L
D000199:Actins / Q000235:genetics*; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D004268:DNA-Binding Proteins / Q000302:isolation & purification / Q000378:metabolism*; D005796:Genes; D006801:Humans; D008969:Molecular Sequence Data; D009132:Muscles / Q000378:metabolism; D009206:Myocardium / Q000378:metabolism; D009210:Myofibrils / Q000378:metabolism*; D009687:Nuclear Proteins / Q000302:isolation & purification / Q000378:metabolism*; D009928:Organ Specificity; D011401:Promoter Regions, Genetic; D012518:Sarcomeres / Q000378:metabolism*
D000199:Actins; D004268:DNA-Binding Proteins; D009687:Nuclear Proteins