Design flexibility in cis-regulatory control of gene expression: synthetic and comparative evidence.

In early Drosophila embryos, the transcription factor Dorsal regulates patterns of gene expression and cell fate specification along the dorsal-ventral axis. How gene expression is produced within the broad lateral domain of the presumptive neurogenic ectoderm is not understood. To investigate transcriptional control during neurogenic ectoderm specification, we examined divergence and function of an embryonic cis-regulatory element controlling the gene short gastrulation (sog). While transcription factor binding sites are not completely conserved, we demonstrate that these sequences are bona fide regulatory elements, despite variable regulatory architecture. Mutation of conserved sequences revealed that putative transcription factor binding sites for Dorsal and Zelda, a ubiquitous maternal transcription factor, are required for proper sog expression. When Zelda and Dorsal sites are paired in a synthetic regulatory element, broad lateral expression results. However, synthetic regulatory elements that contain Dorsal and an additional activator also drive expression throughout the neurogenic ectoderm. Our results suggest that interaction between Dorsal and Zelda drives expression within the presumptive neurogenic ectoderm, but they also demonstrate that regulatory architecture directing expression in this domain is flexible. We propose a model for neurogenic ectoderm specification in which gene regulation occurs at the intersection of temporal and spatial transcription factor inputs.