Periodic Reporting for period 1 - WntELECT (Wnt Enhancer LandscapE CharacTerization)
Reporting period: 2016-05-01 to 2018-04-30
In human breast tumours, deregulation of different Wnt signalling components is frequently observed, ranging from overexpression of individual Wnt genes to elevated levels of the downstream effector beta-catenin. However, in the absence of apparent dominant genetic mutations, the mechanism behind those alterations is unclear. In fact, very little is known about the precise spatio-temporal regulation of Wnt ligands during tissue homeostasis and disease to begin with.
The long-term goal of our research is to uncover how environmental and intrinsic signals are integrated to control dynamic Wnt expression patterns during both normal development and tumour formation. In this project, we aimed to combine bioinformatics, state-of-the-art genome engineering and an innovative screening approach to identify Wnt-associated enhancers in the mammary gland.
We also made important steps towards establishing a high-throughput screening system that will allow the identification of a larger number of active Wnt enhancers. Using a generic CRISPR/Cas9-mediated gene tagging approach, we created clonal cell lines that carry the bright red fluorescent protein mScarlet-I in the last exon of 2 different Wnt genes. Pilot experiments revealed that these cell lines allow us to faithfully measure changes in endogenous Wnt gene expression and will serve as useful tools for studying Wnt signalling biology in development, tissue homeostasis and cancer. Our efforts to optimize the precise screening conditions are still ongoing.