A 13-amino acid amphipathic alpha-helix is required for the functional interaction between the transcriptional repressor Mad1 and mSin3A.
Members of the Mad family of bHLHZip proteins heterodimerize with Max and function to repress the transcriptional and transforming activities of the Myc proto-oncogene. Mad:Max heterodimers repress transcription by recruiting a large multi-protein complex containing the histone deacetylases, HDAC1 and HDAC2, to DNA. The interaction between Mad proteins and HDAC1/2 is mediated by the corepressor mSin3A and requires sequences at the amino terminus of the Mad proteins, termed the SID, for Sin3 interaction domain, and the second of four paired amphipathic alpha-helices (PAH2) in mSin3A. To better understand the requirements for the interaction between the SID and PAH2, we have performed mutagenesis and structural studies on the SID. These studies show that amino acids 8-20 of Mad1 are sufficient for SID:PAH2 interaction. Further, this minimal 13-residue SID peptide forms an amphipathic alpha-helix in solution, and residues on the hydrophobic face of the SID helix are required for interaction with PAH2. Finally, the minimal SID can function as an autonomous and portable repression domain, demonstrating that it is sufficient to target a functional mSin3A/HDAC corepressor complex.
Eilers
A L
AL
Department of Oncological Sciences, Huntsman Cancer Institute, University of Utah, Salt Lake City, Utah 84112-5330, USA.
Billin
A N
AN
Liu
J
J
Ayer
D E
DE
D000595:Amino Acid Sequence; D000818:Animals; D002352:Carrier Proteins; D018797:Cell Cycle Proteins; D002942:Circular Dichroism; D003001:Cloning, Molecular; D017930:Genes, Reporter; D018257:Helix-Loop-Helix Motifs; D006801:Humans; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D009154:Mutation; D009687:Nuclear Proteins / Q000737:chemistry* / Q000235:genetics; D010446:Peptide Fragments / Q000737:chemistry / Q000235:genetics; D010750:Phosphoproteins / Q000737:chemistry* / Q000235:genetics; D017433:Protein Structure, Secondary; D000090063:Proto-Oncogene Mas; D011993:Recombinant Fusion Proteins; D012097:Repressor Proteins / Q000737:chemistry* / Q000235:genetics; D016415:Sequence Alignment; D056264:Sin3 Histone Deacetylase and Corepressor Complex
D002352:Carrier Proteins; D018797:Cell Cycle Proteins; C501384:MAD1L1 protein, human; C000715470:MAS1 protein, human; D046911:Macromolecular Substances; D009687:Nuclear Proteins; D010446:Peptide Fragments; D010750:Phosphoproteins; D000090063:Proto-Oncogene Mas; D011993:Recombinant Fusion Proteins; D012097:Repressor Proteins; C099683:SIN3A transcription factor; D056264:Sin3 Histone Deacetylase and Corepressor Complex