The Athb-1 and -2 HD-Zip domains homodimerize forming complexes of different DNA binding specificities

EMBO J. 1993 Sep;12(9):3507-17.

Abstract

The Arabidopsis Athb-1 and -2 proteins are characterized by the presence of a homeodomain (HD) with a closely linked leucine zipper motif (Zip). We have suggested that the HD-Zip motif could, via dimerization of the leucine zippers, recognize dyad-symmetric DNA sequences. Here we report an analysis of the DNA binding properties of the Athb-1 homeodomain-leucine zipper (HD-Zip-1) domain in vitro. DNA binding analysis performed using random-sequence DNA templates showed that the HD-Zip-1 domain, but not the Athb-1 HD alone, binds to DNA. The HD-Zip-1 domain recognizes a 9 bp dyad-symmetric sequence [CAAT(A/T)ATTG], as determined by selecting high-affinity binding sites from random-sequence DNA. Gel retardation assays demonstrated that the HD-Zip-1 domain binds to DNA as a dimer. Moreover, the analysis of the DNA binding activity of Athb-1 derivatives indicated that a correct spatial relationship between the HD and the Zip is essential for DNA binding. Finally, we determined that the Athb-2 HD-Zip domain recognizes a distinct 9 bp dyad-symmetric sequence [CAAT(G/C)ATTG]. A model of DNA binding by the HD-Zip proteins is proposed.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Arabidopsis / metabolism*
  • Arabidopsis Proteins*
  • Base Sequence
  • DNA / metabolism*
  • DNA-Binding Proteins / biosynthesis
  • DNA-Binding Proteins / metabolism*
  • Glutathione Transferase / metabolism
  • Homeodomain Proteins
  • Leucine Zippers*
  • Macromolecular Substances
  • Models, Structural
  • Molecular Sequence Data
  • Nucleic Acid Conformation
  • Oligodeoxyribonucleotides / metabolism*
  • Plant Proteins / biosynthesis
  • Plant Proteins / metabolism*
  • Polymerase Chain Reaction
  • Protein Biosynthesis
  • Protein Conformation
  • Recombinant Fusion Proteins / metabolism
  • Sequence Homology, Amino Acid
  • Substrate Specificity
  • TATA Box
  • Transcription, Genetic

Substances

  • Arabidopsis Proteins
  • DNA-Binding Proteins
  • HB-2 protein, Arabidopsis
  • Homeodomain Proteins
  • Macromolecular Substances
  • Oligodeoxyribonucleotides
  • Plant Proteins
  • Recombinant Fusion Proteins
  • DNA
  • Glutathione Transferase