The human skeletal alpha-actin gene is regulated by a muscle-specific enhancer that binds three nuclear factors.
The tissue-specific distal promoter of the human skeletal alpha-actin gene (-1282 to -708) induces transcription in myogenic cells approximately 10-fold and, with the most proximal promoter domain (-153 to -87), it synergistically increases transcription 100-fold (Muscat and Kedes 1987). We report here that it is a short fragment of the distal promoter, the distal regulatory element (DRE) from -1282 to -1177 that functions as a muscle-specific, composite enhancer. An internal deletion in the DRE (delta -1282/-1151) in the context of the full-length 2000 bp promoter, resulted in a 10-fold reduction in transcription. Three distinct nuclear proteins, DRF-1, DRF-2, and DRF-3, interact specifically with the DRE between positions -1260 and -1193. A site specific mutation that abolishes DRF-2 binding also results in a 10-fold reduction in transcriptional activity. The DRF-2 nuclear protein has characteristics similar to those of the muscle-specific regulatory factor, MEF-2 (Buskin and Hauschka 1989; Gossett et al., 1989). Like the MEF-2 binding site in the muscle creatine kinase enhancer, the critical DRF-2 binding site is also an A/T-rich sequence element. The DRF-2 nuclear protein binds equally well to the MCK MEF-2 binding site and to the A/T-rich regulatory element of the skeletal muscle fast-twitch troponin C gene (Gahlmann and Kedes 1990). Furthermore, this troponin C site competes in vivo for DRF-2 driven expression of the skeletal alpha-actin gene in C2 cells. The DRF-2 site alone, however, does not activate transcription in muscle cells when linked to the SV40 promoter. We conclude that the DRF-2 binding element is a MEF-2 binding site that is required but insufficient for regulation of muscle-specific skeletal alpha-actin gene expression by the DRE. Thus, muscle-specific regulation of the human skeletal alpha-actin gene appears to require interactions between the other elements of the composite DRE enhancer with the protein:DNA complex formed by DRF-2.
Muscat
G E
GE
Department of Biochemistry, University of Queensland, St. Lucia, Australia.
Perry
S
S
Prentice
H
H
Kedes
L
L
D000199:Actins / Q000235:genetics*; D000818:Animals; D001483:Base Sequence; D002478:Cells, Cultured; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000502:physiology*; D004742:Enhancer Elements, Genetic / Q000502:physiology; D005786:Gene Expression Regulation / Q000502:physiology*; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D009132:Muscles / Q000378:metabolism; D009154:Mutation / Q000502:physiology; D009687:Nuclear Proteins / Q000737:chemistry / Q000502:physiology*; D051381:Rats; D012045:Regulatory Sequences, Nucleic Acid / Q000235:genetics / Q000502:physiology*; D014158:Transcription, Genetic / Q000235:genetics / Q000502:physiology*
D000199:Actins; D004268:DNA-Binding Proteins; D009687:Nuclear Proteins