| Wingless/Wnt family proteins (Wg/Wnt) activates the Wg/Wnt signaling pathway in receiving cells that regulates the expression of its target genes through T cell factor (TCF). The high mobility group (HMG) domain of TCF is known to bind to specific sequences called TCF binding sites (TCF sites) in Wg/Wnt response elements (WREs).;In order to better understand WREs, WREs of naked cuticle (nkd), a direct Wg target, were analyzed. I found that nkd has several WREs which are activated by Wg signaling in multiple tissues, in distinct but overlapping patterns. Further analysis of a nkd-WRE identified a new cis-regulatory element named the Helper site. Helper sites are essential for the activation of several WREs in various fly tissues. As the name implies, this motif potentiates the ability of TCF binding sites to mediate transcriptional activation by Wg signaling. The presence of a Helper site increased the binding affinity of TCF to a classic TCF site in vitro, and the mutation of Helper sites in the WREs reduced recruitment of TCF to the WREs in vivo. This enhanced binding was dependent on the presence of the C-clamp, a domain recently identified to enhance DNA binding of some vertebrate TCF isoforms. Consistently, activation of several WREs containing functional Helper sites was C-clamp dependent.;Interestingly, there is no apparent spacing or orientation requirement for TCF-Helper site pairs. Despite this, we were able to identify two new WREs through a genome-wide search for clusters of TCF-Helper site clusters. Currently, the WREs are beinig been studied in flies by generating transgenic reporter flies.;My data argues that for many targets, DNA binding by the HMG domain of TCF is not sufficient for transcriptional regulation, likely because the protein cannot distinguish WREs from random TCF sites. This problem is overcome by a physical interaction between the C-clamp and the Helper site which increases the affinity of TCF to bona fide WREs. This bipartite recognition of DNA enables TCF to locate and activates WREs. |
