Illuminating transcriptional regulation with genome-wide binding data

Over the last several years, the development of high-throughput in vivo techniques has significantly enhanced our ability to identify targets for transcription factors across the genome. In the first part of this thesis, I use ChIP-on-chip technology to identify targets in the adult mouse liver for Foxa2, a transcription factor that is known to regulate hepatic gene expression. I then introduce an analysis method that demonstrates that Foxa2-containing cis-regulatory modules are not constructed from a random assortment of binding sites for other transcription factors, but rather that the module composition depends on the strength of the Foxa2 consensus site present. Genes containing a cis-regulatory module with a medium or weak Foxa2 consensus site are much more liver-specific than the genes with a strong consensus site. These data show that Foxa2 may have different mechanisms by which it binds to and regulates genes.;ChIP-on-chip is rapidly being replaced by ChIP-Seq technology, which combines ChIP with massively parallel sequencing. Identifying bound regions from the large number of sequence tags produced by ChIP-Seq is a challenging tank. Therefore, in the second part of my thesis, I develop GLITR ( GLobal I&barbelow;dentifier of T&barbelow;arget R&barbelow;egions), an algorithm that accurately identifies enriched regions in target data compared to control data and requires minimal user-defined thresholds. I compare GLITR to several existing methods and show that GLITR has improved sensitivity for identifying bound regions as compared to other algorithms. I also use GLITR to address the issue of sequencing depth, and show that sequencing biological replicates identifies far more binding regions than re-sequencing the same sample. Although ChIP-Seq technology is in its early stages of development, I discuss many of the fundamental aspects of both the experimental and analytical approaches to target identification. The novel methods I present will have a significant impact for future studies that investigate gene regulation by transcription factors.