Analysis of the oligomerization of myogenin and E2A products in vivo using a two-hybrid assay system.
Members of the helix-loop-helix (HLH) family of proteins bind DNA and activate transcription as homo- and heterodimers. Myogenin is a muscle-specific HLH protein that binds DNA in vitro as a heterodimer with several widely expressed HLH proteins, such as the E2A gene products E12 and E47. We describe a method for detection of protein-protein interactions among HLH proteins in vivo in which dimerization through the HLH motif reconstructs a hybrid transcription factor containing the DNA-binding domain of yeast GAL4 linked to one HLH motif and the activation domain of VP-16 linked to another. We have used this assay to investiagate whether myogenin forms homomeric or heteromeric complexes in vivo and to determine whether growth factors and oncogenes that inhibit myogenesis influence myogenin's ability to dimerize. The results show that myogenin heterodimerizes with E12 and E47 in vivo, but it does not homodimerize to a measurable extent. Peptide growth factors, as well as the immediate early gene products c-Jun, v-Fos, and c-Myc, inhibit the activity of myogenin through a mechanism independent of its association with E2A products.
Chakraborty
T
T
Department of Biochemistry and Molecular Biology, University of Texas M. D. Anderson Cancer Center, Houston 77030.
Martin
J F
JF
Olson
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EN
D017456:Adenovirus Early Proteins; D000818:Animals; D002460:Cell Line; D015500:Chloramphenicol O-Acetyltransferase / Q000235:genetics / Q000378:metabolism; D006133:Growth Substances / Q000494:pharmacology; D006367:HeLa Cells; D006801:Humans; D007700:Kinetics; D046911:Macromolecular Substances; D009124:Muscle Proteins / Q000235:genetics / Q000378:metabolism*; D018008:Myogenin; D009856:Oncogene Proteins, Viral / Q000235:genetics / Q000378:metabolism*; D010957:Plasmids; D011994:Recombinant Proteins / Q000378:metabolism; D015534:Trans-Activators / Q000378:metabolism*; D014158:Transcription, Genetic; D015533:Transcriptional Activation; D014162:Transfection
D017456:Adenovirus Early Proteins; D006133:Growth Substances; C503799:MYOG protein, human; D046911:Macromolecular Substances; D009124:Muscle Proteins; D018008:Myogenin; D009856:Oncogene Proteins, Viral; D011994:Recombinant Proteins; D015534:Trans-Activators; D015500:Chloramphenicol O-Acetyltransferase