Abstract
Transcriptional enhancers integrate information derived from transcription factor binding to control gene expression. One key question concerns the extent of trans- and cis-regulatory variation in how co-expressed genes are controlled. The Drosophila CNS midline cells constitute a group of neurons and glia in which expression changes can be readily characterized during specification and differentiation. Using a transgenic approach, we compare the cis-regulation of multiple genes expressed in the Drosophila CNS midline primordium cells, and show that while the expression patterns may appear alike, the target genes are not equivalent in how these common expression patterns are achieved. Some genes utilize a single enhancer that promotes expression in all midline cells, while others utilize multiple enhancers with distinct spatial, temporal, and quantitative contributions. Two regulators, Single-minded and Notch, play key roles in controlling early midline gene expression. While Single-minded is expected to control expression of most, if not all, midline primordium-expressed genes, the role of Notch in directly controlling midline transcription is unknown. Midline primordium expression of the rhomboid gene is dependent on cell signaling by the Notch signaling pathway. Mutational analysis of a rhomboid enhancer reveals at least 5 distinct types of functional cis-control elements, including a binding site for the Notch effector, Suppressor of Hairless. The results suggest a model in which Notch/Suppressor of Hairless levels are insufficient to activate rhomboid expression by itself, but does so in conjunction with additional factors, some of which, including Single-minded, provide midline specificity to Notch activation. Similarly, a midline glial enhancer from the argos gene, which is dependent on EGF/Spitz signaling, is directly regulated by contributions from both Pointed, the EGF transcriptional effector, and Single-minded. In contrast, midline primordium expression of other genes shows a strong dependence on Single-minded and varying combinations of additional transcription factors. Thus, Single-minded directly regulates midline primordium-expressed genes, but in some cases plays a primary role in directing target gene midline expression, and in others provides midline specificity to cell signaling inputs.