Zelda sites activate expression and promote transcription factor binding in a strength-dependent manner. Zeba Wunderlich¹, Rahul Satija², Meghan D. Bragdon¹, Robert K. Bradley³, Angela H. DePace¹. 1) Systems Biology, Harvard Medical School, Boston, MA; 2) Broad Institute of MIT and Harvard, Cambridge, MA; 3) Fred Hutchinson Cancer Research Center, Seattle, WA.

   Genome-wide transcription factor (TF) binding cannot be predicted using TF binding motifs alone. Computational and experimental work has shown that the binding of Zelda, a key TF in the maternal-to-zygotic transition in Drosophila, strongly correlates with the binding of many TFs critical for patterning the embryo. The mechanism by which Zelda affects the binding of other TFs and how binding affects the expression of target genes is currently unknown. We investigate the link between Zelda binding, TF binding and the expression of 4 target genes using ChIP-PCR experiments and quantitative imaging-based measurements of gene expression in blastoderm-stage Drosophila embryos. We created 12 transgenic reporter constructs where lacZ expression is driven by cis-regulatory elements that include Zelda binding sites. For each reporter, we compare the wild-type sequence to variants with the weak, strong, or all Zelda sites deleted. We find that deleting Zelda sites lead to a reduction in expression that depends on the strength of the Zelda sites; deleting only weak sites leads to a slight decrease in reporter expression, while deleting strong sites leads to a larger decrease. ChIP-PCR experiments measuring the change in TF binding and histone modification state are also underway. Together, these experiments will help us understand how Zelda binding affects the binding of other TFs and will help us interpret how changes in Zelda binding sites between individuals and species may lead to changes in gene expression.