Cooperative binding of TEF-1 to repeated GGAATG-related consensus elements with restricted spatial separation and orientation

DNA Cell Biol. 2000 Aug;19(8):507-14. doi: 10.1089/10445490050128430.

Abstract

The human transcriptional enhancer factor (TEF) family includes TEF-1, TEF-3, TEF-4, and TEF-5. The TEFs share a highly conserved 68-amino acid TEA/ATTS DNA-binding domain, which binds to SV40 GT-IIC (GGAATG), SphI (AGTATG), SphII (AGCATG), and muscle-specific M-CAT (GGTATG) enhansons. We determined the optimal DNA-binding consensus sequence for TEF-1. Using a purified GST-TEF-1 fusion protein and a random pool of synthetic oligonucleotides, 31 independent clones were obtained after six rounds of binding site selection. DNA sequences analysis revealed that 16 clones contained direct repeats with a 3-bp spacer (DR3), and 15 clones contained a single binding site. The predominate consensus half-site was GGAATG (67%), and the other elements were of the form G(A)GA(T/C)ATG. The TEF-1 bound to the DR3 as a dimer in a cooperative manner. Cooperative binding was dependent on the spacing and orientation of the half-sites and was inhibited by deoxycholate treatment, providing evidence that protein-protein interactions were involved. The data suggest that TEF dimerization is important for its ability to modulate gene transcription.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Base Sequence
  • Binding Sites
  • Consensus Sequence*
  • DNA-Binding Proteins / genetics
  • DNA-Binding Proteins / metabolism*
  • Humans
  • Nuclear Proteins*
  • Protein Binding
  • Recombinant Fusion Proteins / metabolism
  • Repetitive Sequences, Nucleic Acid*
  • TEA Domain Transcription Factors
  • Transcription Factors / genetics
  • Transcription Factors / metabolism*

Substances

  • DNA-Binding Proteins
  • Nuclear Proteins
  • Recombinant Fusion Proteins
  • TEA Domain Transcription Factors
  • TEAD1 protein, human
  • Transcription Factors