Hedgehog Signaling Coordinates Stochastic and Stereotyped Patterns in the Drosophila Eye

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Project Summary Development of an organism requires both stereotyped and stochastic patterning. Stereotyped patterning robustly generates nearly identical structures across individuals. In contrast, stochastic cell fate specification produces randomized patterns that are unique to each individual. Stochastic fate decisions are required for the development of many sensory organs, including visual and olfactory systems. Despite their importance, how the molecular mechanisms controlling stereotyped and random patterns intersect within the same tissue has not been addressed. This project aims to determine how gene regulatory mechanisms are tuned to generate highly regular patterns and stochastic patterns in the same tissue using the Drosophila eye as a model. The Drosophila eye is composed of ~800 ommatidia in a near perfect array. Each ommatidium comprises eight photoreceptors (R1-8) which develop in a predictable fashion. As photoreceptors are differentiating during larval eye development, a wave of morphogenesis driven by Hedgehog (Hh) signaling drives the highly reproducible structure of the eye. Underlying the uniform morphology of the fly eye is a random pattern of photoreceptor subtypes. Two R7 photoreceptor subtypes are defined by expression of light-detecting Rhodopsin proteins. Random patterning of these two R7 subtypes is controlled by stochastic ON/OFF expression of the transcription factor, Spineless (Ss). SsON R7s express Rhodopsin 4 (Rh4), whereas SsOFF R7s express Rhodopsin 3 (Rh3). ss is regulated by an interplay of transcription and chromatin regulation during larval eye development. I found that Hh signaling plays a second role in eye development to regulate stochastic patterning. hh mutants display a reduction in the percentage of SsON R7s. Cubitus Interruptus (Ci), an effector of Hh signaling, binds at an eye specific enhancer in ss. This site overlaps with a binding site for Klumpfuss (Klu), a repressor of ss, suggesting competitive binding and regulation. I hypothesize that Hh signaling is finely tuned to drive stereotyped eye patterning and induce ss transcription in precursors to generate stochastic R7 subtype patterning. I will test this hypothesis by 1) Determining how the Hh pathway regulates stochastic ss expression, 2) Describing how antagonism between Ci and Klu regulates ss expression, and 3) Determining how Hh signaling and chromatin regulation are integrated at the ss locus. Together, these experiments will provide the first analysis of coordination between stochastic and stereotyped gene expression within a single tissue and will enhance our mechanistic understanding of stochastic cell fate specification.