Co-operativity of functional domains in the muscle-specific transcription factor Myf-5.
Myf-5 is a member of a family of muscle-specific transcription factors that activate myogenesis in 10T1/2 fibroblasts. Here we report the analysis of Myf-5 structural domains that are responsible for its biological activity. Site-directed mutagenesis revealed that two clusters of basic amino acids within a conserved basic region and two amphipathic helices within the adjacent HLH domain are essential for sequence-specific DNA binding and hetero-oligomerization, respectively. Transcriptional activation by Myf-5 requires two additional domains located in the amino- and carboxyl-termini. The two domains apparently co-operate since deletion of either one results in inactivation. Chimeric proteins between DNA binding domain of the yeast transcription factor GAL4 and the separate Myf-5 transactivator domains exhibit activity that is enhanced when both regions are combined. Dimerization of Myf-5 with the ubiquitously expressed bHLH protein E12 not only increases the affinity for DNA but also stimulates transactivation independently of DNA binding. The Myf-5 transactivator domains are dependent for activity on a specific amino acid sequence motif within the basic region when Myf-5 activity is mediated through the E-box DNA recognition sequence but not when DNA binding occurs through the GAL4 DNA binding domain. This demonstrates that muscle-specific transactivation by Myf-5 requires the collaboration of two activation domains and the DNA binding region in addition to sequence-specific DNA binding. Transcriptional activation and interaction with DNA are executed by separable domains; however, transactivation is influenced by the basic region in a manner distinguishable from DNA binding.
Winter
B
B
Department of Toxicology, University of Hamburg, Medical School, FRG.
Braun
T
T
Arnold
H H
HH
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002478:Cells, Cultured; D004247:DNA / Q000235:genetics; D004268:DNA-Binding Proteins / Q000378:metabolism; D005656:Fungal Proteins / Q000378:metabolism; D051379:Mice; D008969:Molecular Sequence Data; D009124:Muscle Proteins / Q000235:genetics / Q000378:metabolism*; D009132:Muscles / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed; D051787:Myogenic Regulatory Factor 5; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins; D015534:Trans-Activators; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D014158:Transcription, Genetic; D015533:Transcriptional Activation
D004268:DNA-Binding Proteins; D005656:Fungal Proteins; C052130:GAL4 protein, S cerevisiae; D009124:Muscle Proteins; C497418:Myf5 protein, mouse; D051787:Myogenic Regulatory Factor 5; D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins; D015534:Trans-Activators; D014157:Transcription Factors; D004247:DNA