Structures of the Ets Protein DNA-binding Domains of Transcription Factors Etv1, Etv4, Etv5, and Fev: DETERMINANTS OF DNA BINDING AND REDOX REGULATION BY DISULFIDE BOND FORMATION

J Biol Chem. 2015 May 29;290(22):13692-709. doi: 10.1074/jbc.M115.646737. Epub 2015 Apr 12.

Abstract

Ets transcription factors, which share the conserved Ets DNA-binding domain, number nearly 30 members in humans and are particularly involved in developmental processes. Their deregulation following changes in expression, transcriptional activity, or by chromosomal translocation plays a critical role in carcinogenesis. Ets DNA binding, selectivity, and regulation have been extensively studied; however, questions still arise regarding binding specificity outside the core GGA recognition sequence and the mode of action of Ets post-translational modifications. Here, we report the crystal structures of Etv1, Etv4, Etv5, and Fev, alone and in complex with DNA. We identify previously unrecognized features of the protein-DNA interface. Interactions with the DNA backbone account for most of the binding affinity. We describe a highly coordinated network of water molecules acting in base selection upstream of the GGAA core and the structural features that may account for discrimination against methylated cytidine residues. Unexpectedly, all proteins crystallized as disulfide-linked dimers, exhibiting a novel interface (distant to the DNA recognition helix). Homodimers of Etv1, Etv4, and Etv5 could be reduced to monomers, leading to a 40-200-fold increase in DNA binding affinity. Hence, we present the first indication of a redox-dependent regulatory mechanism that may control the activity of this subset of oncogenic Ets transcription factors.

Keywords: DNA methylation; ETS transcription factor family; dimerization; protein structure; protein-DNA interaction; redox regulation.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adenovirus E1A Proteins / chemistry*
  • Amino Acid Sequence
  • Chromatography, Liquid
  • DNA / chemistry
  • DNA-Binding Proteins / chemistry*
  • Disulfides / chemistry*
  • Gene Expression Regulation
  • Humans
  • Models, Molecular
  • Molecular Sequence Data
  • Mutagenesis, Site-Directed
  • Neoplasms / metabolism
  • Nuclear Proteins / chemistry*
  • Oxidation-Reduction
  • Oxygen / chemistry
  • Protein Binding
  • Protein Conformation
  • Protein Multimerization
  • Proto-Oncogene Proteins / chemistry*
  • Proto-Oncogene Proteins c-ets / chemistry*
  • Recombinant Proteins / chemistry
  • Sequence Homology, Amino Acid
  • Spectrometry, Mass, Electrospray Ionization
  • Transcription Factors / chemistry*

Substances

  • Adenovirus E1A Proteins
  • DNA-Binding Proteins
  • Disulfides
  • ETV1 protein, human
  • ETV4 protein, human
  • ETV5 protein, human
  • FEV protein, human
  • Nuclear Proteins
  • Proto-Oncogene Proteins
  • Proto-Oncogene Proteins c-ets
  • Recombinant Proteins
  • Transcription Factors
  • DNA
  • Oxygen

Associated data

  • PDB/2PYR
  • PDB/3ZP5
  • PDB/4AVP
  • PDB/4BNC
  • PDB/4CO8
  • PDB/4UNO
  • PDB/4UUV