In the past decade, the genomes of many organisms have been completely sequenced. Two striking observations are that the genomes of higher eukaryotes contain significantly fewer genes than originally predicted, and that most of the genomic DNA has, as yet, no recognized function. The vastly increased biological complexity of these species must therefore be critically dependent on very tightly directed expression of gene networks during their development. Much of this complex regulation depends upon uncharacterized functions of non-protein coding intergenic regions. A central question is the molecular mechanism by which enhancers are recruited to specific gene promoters during embryonic development. An exciting recent advance is the investigators' identification of a novel DNA sequence which directly regulates promoter-enhancer interactions. The studies proposed here will specifically address the in vivo functional role of this newly characterized cis-regulatory element through transgenic and molecular experimental approaches. ? ? Regulation of homeotic (Hox) gene expression from the bithorax complex (BX-C) in Drosophila melanogaster will be the focus of this application. The correct spatial and temporal expression of the three protein-coding Hox genes from the BX-C is essential for the control of developmental cell fate along the anterior-posterior axis of the embryo. The BX-C also provides some of the best current evidence for the function of intergenic, non-coding DNA in regulation of gene expression. The regulatory regions providing this exquisite control are dispersed throughout the 150 kb of DNA that defines the BX-C. The overall scientific goal will be to elucidate the molecular mechanisms by which these regulatory regions achieve coordinated control of gene expression across the entire BX-C. The major focus will be to define regulatory mechanisms that control specific promoter-enhancer interactions in the BX-C. The proposed studies will provide major advances in our understanding of how the genome utilizes epigenetic and cis-regulatory mechanisms to provide the precise gene expression patterns critical to development in multicellular organisms. Further, given the remarkable conservation of Hox gene complexes across evolution, the findings will provide broad insights into developmental regulation across all evolutionary orders. Consequently, these studies will have a considerable impact on our comprehension of a number of human developmental defects associated with misexpression of Hox genes during embryonic development. ? ? ?
