The objective of the research proposed is to address a fundamental question in developmental biology: what are the molecular mechanisms that direct pattern formation in developing animal appendages? Critical to the generation of exquisitely patterned body plans of multicellular organisms is position-specific gene expression during development. The Drosophila adult leg provides a simple model system to investigate this process. On the leg, a group of small mechanosensory bristles are organized into a series of longitudinal rows, a pattern which depends largely on the periodic expression of just two genes: the antinueral gene, hairy (h) and the proneural gene, achaete (ac). In pupal legs, h is expressed in two pairs of longitudinal stripes, one pair that traverses the dorsal/ventral (D/V) axis (D/V-h) and another pair that runs along the anterior/poster (A/P) axis (A/P-h). A stripe of ac expression on either side of each h domain defines the primordia of the leg bristle rows. In the absence of h function, ac expression expands into the regions normally occupied by H, resulting in disorganized bristle rows in the adult. Hence, mechanistic insight into the periodic patterning of leg mCs can be gained by investigating the regulation of h and ac expression.
The focus of the studies proposed will be on the regulation D/V-h expression during leg imaginal disc development. Regulation of D/V-h expression is surprisingly complex and is directed by dorsal (D-h) and ventral (V-h) specific enhancers that integrate signals from the Hedgehog (Hh), Decapentaplegic (Dpp), Wingless (Wg), and the Epidermal growth factor receptor (EGFR) pathways. The D-h enhancer consists of two subelements, a Hh responsive element (D-h-HE), which activates D-h expression in a broad domain adjacent to the A/P boundary of the leg disc, and a repression element (D-h-RE) which functions to refine D-h expression along the A/P and D/V axes. Preliminary studies suggest that the transcriptional repressor Brinker (Brk) acts through the D-h-RE to repress D-h-HE directed expression. This observation suggests a novel role for Brk in repressing expression directed by a Hh responsive enhancer element. Furthermore, Dpp and EFGR signal through the D-h-RE to block Brk repression of D-h expression. V-h expression, on the other hand, is controlled by the combined input of Hh and Wg. Correspondingly, potential Hh and Wg response sites have been identified in the V-h enhancer.
Genetic and molecular analysis of the D and V-h enhancers will be extended with the goal of broadening the understanding of the molecular mechanisms involved in signal integration and interactions between activators and repressors. These studies will provide further insight into the relationship between enhancer function and pattern formation. The specific aims are to: 1. To determine how the D-h subelements, the D-h-HE and D-h-RE function coordinately to integrate the Hh, Dpp and EGFR signals. 2. To identify functional sequences in the V-h enhancer and trans-acting factors that control spatial and temporal expression of V-h.
The research proposed will take place at the University of Illinois at Chicago (UIC). UIC is an urban campus with a diverse undergraduate and graduate population. Both minority and nonminority undergraduate and graduate students have particiapted in ongoing research in the P.I.'s laboratory. Other efforts to broaden the impact of research efforts: 1. introduction of Drosophila as model organism for development studies to students in the laboratory section of an undergraduate Developmental Biology course and 2. yearly visits to two local elementary schools to make presentations on Drosophila genetics and introduction to DNA. 3. judging science fair projects for local elementary and high schools.
