During development of a multicellular animal, cell proliferation must be orchestrated with cell differentiation for proper tissue formation. Cells coordinate their activities by segregating into separate domains - or compartments - and establishing regions of reduced cell mixing at their boundaries. Cell fate boundaries organize patterning and morphogenesis of a wide range of tissues, including the eye equator and wing margin in Drosophila, and the apical ectodermal ridge, somite and hindbrain rhombomeres in vertebrates. Little is known about the mechanisms that generate cell fate boundaries and translate compartment boundaries into the structural features of mature tissue.
The Drosophila bunched gene, a member of the highly conserved TSC-22/GILZ family of transcription factors, establishes an epithelial cell fate boundary during oogenesis. bunched, which encodes three distinct protein isoforms whose structural features correspond to three isoforms of the mouse TSC-22 gene, is regulated by long-range morphogens to set the position of a cell fate boundary. The Bunched1 isoform regulates cell affinities and short-range Notch signaling at a cell fate boundary that coincides with a key structural feature of the fly eggshell, the collar of the operculum.
Using a simple model tissue and applying powerful molecular and genetic approaches, the goals of this proposal are to: (1) determine how long-range morphogen signaling is refined to short-range Notch activation via Bunched1; and (2) characterize how interactions among the three Bunched isoforms pattern cell fates. The long term goal of this proposal is to elucidate the complex, conserved mechanism by which cell fate boundaries are established.
Broader Impact A fundamental question in development is how tissue subdivision occurs by genes, initially expressed in broad, overlapping domains, which regulate the expression of downstream genes to establish a defined pattern. The TSC-22 family of proteins is widely conserved in animals but extensive mammalian tissue culture studies have shed no light on their developmental function.
Experiments proposed will give basic insights into how Bunched protein isoforms interact to regulate gene transcription at forming cell fate boundaries and illuminate similar regulatory processes in limbs, nervous system and other vertebrate and invertebrate tissues. Experiments proposed are designed to integrate research with a program of teaching and training undergraduate and graduate students. The focus on a Drosophila gene similar to human TSC-22, a gene associated with several human cancers and diabetic neuropathy, will convey to students the potential of model organisms to elucidate developmental mechanisms as a basis to understand human disease. The logical series of experiments detailed in these Aims are divisible into larger projects suitable for graduate students and smaller projects appropriate for undergraduates. Work outlined here will complement undergraduate research, including scholars programs for extended research and summer research (SEARCH and Sapere Vedere scholarships) and the Ph.D. training program. This project will enhance teaching: (1) Developmental Biology for undergraduates and Advanced Cell Biology, a graduate level course emphasizing the use of animal model systems like Drosophila, C. elegans, and yeast as genomic tools. This work will further development of a lab course for Developmental Biology, based in part on the genetic experiments planned here.
