The development of a complex organ such as the eye involves the specification of multiple distinct cell types and the integration of their functions. In Drosophila, the light-detecting photoreceptors and the glial-like cone and pigment cells that secrete the lens and produce screening pigments arise from a single field of equivalent progenitors. Commitment to one of these cell fates requires extracellular signals to be integrated with a complex network of intrinsic transcription factors. Although receptor tyrosine kinase signaling has been known for many years to induce both photoreceptors and cone cells, most of its target genes remain unknown. It is also unclear which intrinsic transcription factors distinguish these cell identities. The zinc finger transcription factor Glass had been thought to specify the photoreceptor fate, but our recent work showed that it acts in each of the three cell types to promote their normal differentiation. This proposal seeks to understand how these common factors activate distinct differentiation pathways.
The first aim will look for direct target genes of receptor tyrosine kinase signaling and Glass during retinal differentiation. Targeted DNA adenine methyltransferase identification (DamID) will be used to identify direct target genes of Pointed, the transcription factor that mediates Epidermal growth factor receptor (EGFR) signaling, in undifferentiated cells, photoreceptors and cone cells. A similar approach will be used to identify Glass target genes, and both datasets will be compared to transcriptomic analysis of genes that change their expression in Egfr or glass mutants. The goal of these experiments is to identify transcription factors that are induced by EGFR signaling to promote the differentiation of photoreceptors and cone cells, and elucidate how Glass feeds into their regulation.
The second aim concerns how transcriptional repressors restrict the effects of Glass to drive cell type-specific gene expression. Two defined regulatory regions that are known to drive photoreceptor-specific expression in a Glass-dependent manner will be used to identify repressors that prevent Glass from activating these genes in cone and pigment cells. The importance of these repressor binding sites will be tested in the endogenous genomic context. In a complementary approach, enhancer regions that drive Glass-dependent expression specifically in cone or pigment cells will be identified and their regulation characterized. In combination, these experiments will reveal how distinct cell identities can be specified from common progenitors using a limited set of signals and transcription factors, a process that also occurs in the mammalian retina. The results will help to refine our ability to produce specific cell types from stem cells for regenerative medicine.
Organs are built from multiple cell types that must be specified and integrated into a functional unit. In order to understand how cells choose their fates and develop different functional capacities, this proposal will determine how interactions between signaling pathways and transcriptional networks promote the differentiation of all the cell types of the Drosophila eye. The goal is to uncover mechanisms that will both explain genetic diseases of the eye, and improve our ability to design cell replacement therapies.
