The Hox-1.3 homeo box protein is a sequence-specific DNA-binding phosphoprotein

We report that the murine Hox-1.3 homeo domain protein is a nuclear phosphoprotein capable of binding to specific DNA sequences. DNase I protection of the Hox-1.3 gene promoter region with the Hox-1.3 protein identifies a binding site 144 bp upstream from the start of transcription. Both phosphorylated and nonphosphorylated forms bind DNA directly in a sequence-specific manner. Electrophoretic mobility shift assays were performed with a set of synthetic oligonucleotides representing either the DNase I-protected region of the Hox-1.3 gene or partially homologous sequences present in promoter regions of other characterized viral, yeast, and mammalian genes. From the results, we deduce a consensus binding motif of CPyPyNATTAT/GPy. Base substitutions in the core ATTA sequence severely reduce or abolish binding. In the SV40 enhancer, the Hox-1.3 binding motif overlaps both the octamer (Octa2) and the transactivator protein-1 (AP-1) binding sites. The Hox-1.3 binding motif also overlaps the nuclear factor III (NF-III) octamer motif in the adenovirus-2 origin of DNA replication. Overlap among DNA-binding sites suggests that regulation imparted by certain cis-elements may be integrated by these different factors.