Collaborative interactions between MEF-2 and Sp1 in muscle-specific gene regulation.
Previous investigations have demonstrated synergistic interactions in vivo between CCAC and A/T-rich nucleotide sequence motifs as functional components of muscle-specific transcriptional enhancers. Using CCAC and A/T-rich elements from the myoglobin and muscle creatine kinase (MCK) gene enhancers, Sp1 and myocyte-specific enhancer factor-2 (MEF-2) were identified as cognate binding proteins that recognize these sites. Physical interactions between Sp1 and MEF-2 were demonstrated by immunological detection of both proteins in DNA binding complexes formed in vitro by nuclear extracts in the presence of only the A/T sequence motif, by coprecipitation of recombinant MEF-2 in the presence of a glutathione-S-transferase-Sp1 fusion protein bound to glutathione beads, and by a two-hybrid assay in Saccharomyces cerevisiae. The interaction with Sp1 in vitro and in vivo is specific for MEF-2 and was not observed with serum response factor, a related MADS domain protein. Forced expression of Sp1 and MEF-2 in insect cells otherwise lacking these factors promotes synergistic transcriptional activation of a promoter containing binding sites for both proteins. These data expand the repertoire of functional and physical interactions between lineage-restricted (MEF-2) and ubiquitous (Sp1) transcription factors that may be important for myogenic differentiation.
Department of Internal Medicine, University of Texas Southwestern Medical Center, Dallas 75235-8573, USA.
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J
Grayson
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Bassel-Duby
R S
RS
Williams
D000818:Animals; D002460:Cell Line; D003402:Creatine Kinase / Q000235:genetics*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D029721:Drosophila Proteins; D004331:Drosophila melanogaster; D004742:Enhancer Elements, Genetic; D005786:Gene Expression Regulation; D005982:Glutathione Transferase; D008156:Luciferases; D064326:MEF2 Transcription Factors; D051379:Mice; D009132:Muscles / Q000378:metabolism*; D018006:Myogenic Regulatory Factors; D009211:Myoglobin / Q000235:genetics; D009687:Nuclear Proteins / Q000378:metabolism; D009928:Organ Specificity; D014176:Protein Biosynthesis; D011993:Recombinant Fusion Proteins / Q000302:isolation & purification / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics; D026362:Serum Response Factor; D016329:Sp1 Transcription Factor / Q000378:metabolism*; D014157:Transcription Factors / Q000378:metabolism*; D014162:Transfection
D004268:DNA-Binding Proteins; D029721:Drosophila Proteins; D064326:MEF2 Transcription Factors; C499409:Mef2 protein, Drosophila; D018006:Myogenic Regulatory Factors; D009211:Myoglobin; D009687:Nuclear Proteins; D011993:Recombinant Fusion Proteins; D026362:Serum Response Factor; D016329:Sp1 Transcription Factor; D014157:Transcription Factors; D008156:Luciferases; D005982:Glutathione Transferase; D003402:Creatine Kinase