Mapping activation and repression domains of the vnd/NK-2 homeodomain protein

A transient transfection assay using Drosophila S2 tissue culture cells and WT and mutant Drosophila vnd/NK-2 homeobox cDNAs was used to localize repression and activation domains of vnd/NK-2 homeodomain protein. A repression domain was identified near the N terminus of vnd/NK-2 homeodomain protein (amino acid residues 154-193), which contains many hydrophobic amino acid residues. The major determinants of the repression domain were shown to be amino acid residues F155, W158, I161, L162, L163, and W166. Truncated protein consisting of the N-terminal repression domain and the DNA-binding homeodomain repressed transcription as efficiently as WT vnd/NK-2 protein. An activation domain was identified between the tinman domain and the homeodomain (amino acid residues 277-543), which consists of a glutamine-rich subdomain and two acidic subdomains. No effect was detected of the tinman domain or the NK-2-specific domain on either activation or repression of a beta-galactosidase reporter gene.