EXPLORING THE HOMEOBOX

In Drosophila, homeotic mutations lead to the transformation of structures of one body segment into the corresponding structures of another segment. These mutations identify master regulator genes which specify segmental identity along the antero-posterior body axis. Dominant gain and recessive loss-of-function mutations generate to opposite segmental transformations. The cloning of the homeotic Antennapedia (Antp) gene led to the discovery of the homeobox, a 180-bp DNA segment characteristic for homeotic genes. It encodes the DNA-binding domain of the respective proteins which was designated as the homeodomain. Homeodomain proteins are transcriptional regulators which specify the body plan by controlling the transcription of their subordinate target genes. By inserting the Antp cDNA into a heat-inducible expression vector, the body plan can be altered in a predictable way. Using the homeobox as a probe, homologous Hox genes from vertebrates have been cloned. In the mouse, dominant gain and recessive loss-of-function mutations result in segmental transformations of opposite direction, as in Drosophila. Also, the mouse Hox genes can partially substitute the homologous Drosophila genes in transgenic flies. Therefore, the genetic control of the body plan is much more universal than anticipated. The three-dimensional structure of the Antp homeodomain and its complex with a consensus DNA-binding site was determined by nuclear magnetic resonance (NMR) spectroscopy. The homeodomain essentially consists of four alpha-helices, a helix-turn-helix motif, and a flexible N-terminal arm. Base-specific contacts are made by both the recognition helix and the N-terminal arm. The essential features of the NMR model are consistent with in vivo data obtained by mutations altering the DNA binding and the functional specificity in transgenic flies.