In vivo footprinting of a muscle specific enhancer by ligation mediated PCR.
In vivo protein-DNA interactions at the developmentally regulated enhancer of the mouse muscle creatine kinase (MCK) gene were examined by a newly developed polymerase chain reaction (PCR) footprinting procedure. This ligation mediated, single-sided PCR technique permits the exponential amplification of an entire sequence ladder. Several footprints were detected in terminally differentiated muscle cells where the MCK gene is actively transcribed. None were observed in myogenic cells prior to differentiation or in nonmuscle cells. Two footprints appear to correspond to sites that can bind the myogenic regulator MyoD1 in vitro, whereas two others represent muscle specific use of apparently general factors. Because MyoD1 is synthesized by undifferentiated myoblasts, these data imply that additional regulatory mechanisms must restrict the interaction between this protein and its target site prior to differentiation.
Mueller
P R
PR
Division of Biology, California Institute of Technology, Pasadena 91125.
Wold
B
B
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D002460:Cell Line; D003402:Creatine Kinase / Q000235:genetics*; D004247:DNA / Q000032:analysis* / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D004742:Enhancer Elements, Genetic; D005784:Gene Amplification; D015870:Gene Expression; D005809:Genes, Regulator; D051379:Mice; D008969:Molecular Sequence Data; D009132:Muscles / Q000201:enzymology*; D016133:Polymerase Chain Reaction; D011485:Protein Binding; D011499:Protein Processing, Post-Translational; D013698:Templates, Genetic; D050656:Transcription Factor AP-2; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism
D004268:DNA-Binding Proteins; D050656:Transcription Factor AP-2; D014157:Transcription Factors; D004247:DNA; D003402:Creatine Kinase