The role of FOXK transcription factors in Wnt signal transduction

Wnt signalling is a highly conserved signal transduction pathway with key functions in developmental processes including patterning during embryogenesis and stem cell homeostasis in the adult organism. Wnt signals are received at the cell membrane and transduced through cytoplasmic and nuclear multiprotein complexes; the degradasome, signalosome and enhanceosome. The latter is tethered to enhancers by T-cell transcription factors (TCF), and core components include the chromatin reader Pygopus as well as the transcriptional repressor Groucho/TLE3. While much is known about the basic interactions within the enhanceosome, it remains elusive how the enhanceosome accesses native chromatin and is established during development. Furthermore, it is controversial how transcriptional inhibition is re-installed after active signalling ceases and whether Groucho/TLE3 remains attached to the enhanceosome throughout signalling. In order to elucidate novel binding partners, as well as assess potential restructuring of the enhanceosome during different stages of signalling, I employed a proteomics approach based on proximity labelling (BioID) using TLE3 and Pygopus as bait proteins. This revealed little changes of the core enhanceosome subunits between active or inactive signalling but indicated the Forkheadbox transcription factors FOXK1/2 as potential members of the Wnt enhanceosome. FOXK proteins are interesting candidates as they are thought to function as pioneer factors that bind to closed chromatin resulting in its opening and increased accessibility for transcription factors. Use of a minimal Ubx enhancer system corroborates the function of FOX proteins in general during Wg signalling in the fly. I found that a previously published FoxK fly mutant does not carry a FoxK null allele and consequently generated new knockout alleles using CRISPR/Cas9. Phenotypic analysis identified a function for FoxK during early embryogenesis whereby null mutants derived from germ line clones arrest in early development and strikingly show lack of Wg as well as segmentation and gastrulation defects. I identified a specific requirement of FoxK for patterning of the wing blade. I further assessed a putative interaction of FOXK1/2 with DVL, a key player of the signalosome. Contrary to published results, I was able to show that FOXK1/2 do not function to recruit DVL into the nucleus and do not drive expression of a TCF reporter. This work expands our knowledge on FOXK transcription factors during fly development and in mammalian cells and establishes a function for FOX proteins during processes known to be regulated by Wg.

Name dc:type.qualificationname

Doctor of Philosophy (PhD)

Level dc:type.qualificationlevel

Doctoral

Grantor dc:publisher.institution

University of Cambridge

Year dc:date.issued

2020

Author dc:creator

• Heinke, Lisa

Advisor dc:contributor.advisor

• Bienz, Mariann

• Molecular Biology
• Wnt signalling

Licence

• https://apollo8-f-pro.lib.cam.ac.uk/bitstreams/a8f07694-5631-41c1-897a-3f93ce1e9585/download
• https://www.rioxx.net/licenses/all-rights-reserved/

Language dc:language

eng

DOI dc:identifier.doi

https://doi.org/10.17863/CAM.58222

OAI identifier oai:identifier

oai:www.repository.cam.ac.uk:1810/311134