PBX PROTEINS DISPLAY HEXAPEPTIDE-DEPENDENT COOPERATIVE DNA-BINDING WITH A SUBSET OF HOX PROTEINS

The human proto-oncogene PBX1 codes for a homolog of Drosophila extrad enticle, a divergent homeo domain protein that modulates the developme ntal and DNA-binding specificity of select HOM proteins. We demonstrat e that wild-type Pbx proteins and chimeric E2a-Pbx1 oncoproteins coope ratively bind a consensus DNA probe with HoxB4, B6, and B7 of the Ante nnapedia class of Hox/HOM proteins. Specificity of Hox-Pbx interaction s was suggested by the inability of Pbx proteins to cooperatively bind the synthetic DNA target with HoxA10 or Drosophila even-skipped. Site -directed mutagenesis showed that the hexapeptide motif (IYPWMK) upstr eam of the Hox homeo domain was essential for HoxB6 and B7 to cooperat ively bind DNA with Pbx proteins. Engraftment of the HoxB7 hexapeptide onto HoxA10 endowed it with robust cooperative properties, demonstrat ing a functional role for the highly conserved hexapeptide element as one of the molecular determinants delimiting Hox-Pbx cooperativity. Th e Pbx homeo domain was necessary but not sufficient for cooperativity, which required conserved amino acids carboxy-terminal of the homeo do main. These findings demonstrate that interactions between Hox and Pbx proteins modulate their DNA-binding properties, suggesting that Pbx a nd Hox proteins act in parallel as heterotypic complexes to regulate e xpression of specific subordinate genes.