A STRUCTURAL MODEL FOR A HOMEOTIC PROTEIN-EXTRADENTICLE-DNA COMPLEX ACCOUNTS FOR THE CHOICE OF HOX PROTEIN IN THE HETERODIMER

The genes of the homeotic complex (HOX) encode DNA binding homeodomain proteins that control developmental fates by differentially regulatin g the transcription of downstream target genes. Despite their unique i n vivo functions, disparate HOX proteins often bind to very similar DN A sequences in vitro. Thus, a critical question is how HOX proteins se lect the correct sets of target genes in vivo. The homeodomain protein s encoded by the Drosophila extradenticle gene and its mammalian homol ogues, the pbx genes, contribute to HOX specificity by cooperatively b inding to DNA with HOX proteins. For example, the HOX protein labial c ooperatively binds with extradenticle protein to a 20-bp oligonucleoti de that is sufficient to direct a labial-like expression pattern in Dr osophila embryos. Here we have analyzed the protein-DNA interactions t hat are important for forming the labial-extradenticle-DNA complex. Th e data suggest a model in which labial and extradenticle, separated by only 4 bp, bind this DNA as a heterodimer in a head-to-tail orientati on. We have confirmed several aspects of this model by characterizing extradenticle-HOX binding to mutant oligonucleotides. Most importantly , mutations in base pairs predicted to contact the HOX N-terminal arm resulted in a change in HOX preference in the heterodimer, from labial to Ultrabithorax. These results demonstrate that extradenticle prefer s to bind cooperatively with different HOX proteins depending on subtl e differences in the heterodimer binding site.