Measuring and Modeling Enhancers in Perturbed Drosophila Melanogaster Embryos

Abstract

The diversity of animal shapes and sizes, colors and textures, or behaviors and habitats all depend on specialized cells. A newly fertilized embryo must build all these specialized cell types, a process called differentiation. Much of differentiation depends on appropriately turning genes on and off in each cell type. Cell type specific control of gene expression is encoded in a type of regulatory DNA called enhancers. I am interested in how enhancers control the cell type specific gene expression that enables specialized cell functions.

Enhancers read in information from regulatory proteins and output a level of gene expression. This conversion from input regulator concentrations to output expression level is a computation. I use quantitative measurement and computational modeling to study how enhancers compute. In embryos, many regulatory proteins bind to enhancers, and some will turn an enhancer on, while others will turn it off. This complex process is greatly simplified by employing computational models. These models can test whether all regulators have been identified (and if not, find the missing ones) and quantify the relationships between regulators. The relationships between regulators reflect the underlying molecular mechanisms used in the cell; when several models can fit the data, perturbation experiments can be used to distinguish the models and underlying mechanisms. However, most computational models of gene expression in animals have not been rigorously validated by perturbation experiments. A major contribution of my thesis work was developing methods for testing models.

To test computational models for how enhancers compute gene expression patterns, I experimentally manipulated the concentrations of regulatory proteins and precisely measured output gene expression patterns. Using the Drosophila melanogaster blastoderm embryo, I first developed efficient and scalable techniques for making perturbations to regulatory protein concentrations. This technique revealed a postulated property of development: that embryos mitigate the impact of perturbations by preventing the creation of new cell types. I then used two perturbations to test computational models of an enhancer, finding they were incomplete and discovering new regulatory connections. My work illustrates how computational modeling and quantitative measurement are powerful tools for untangling how regulatory DNA operates in embryos.