Hohenstein, Peter

Wood, Andrew

Krishna, Anagha

2019-07-17T12:13:10Z

2019-07-17T12:13:10Z

2019-06-29

Canonical Wnt signaling is one of the most extensively studied signaling cascades, with a multifunctional role in development and disease. Activating mutations in β-catenin, the key regulator of this pathway, have been observed in many types of cancer. The general hypothesis is that, all these activating mutations in β-catenin affect the protein turnover and localization, and thus aberrantly activate the Wnt target genes that are capable of modulating multiple aspects of tumourigenesis. Although various residues in β-catenin were found to be mutated in different cancer types, a detailed analysis of the individual mutations has been lacking so far. In this project, firstly I sought to explore the frequency and distribution of mutations in the β-catenin gene across different cancer types using the COSMIC (Catalogue of Somatic Mutations in Cancer) database. The analysis of the β-catenin mutational spectrum revealed a preferential selection of different residues and amino acid substitutions among the different cancer types. This specific selectivity of mutations indicated a difference in phenotypic effect from different β-catenin mutations. Furthermore, through "saturation mutagenesis" of the β-catenin hotspot region (L31-G50) and by generating independent clonal cell lines harbouring various substitution mutations at the selected top six β-catenin residues through "multiplex targeting", I tried to understand whether or not there exists a genotype-phenotype correlation among the β-catenin mutations. Using these complementary experimental approaches, I convincingly demonstrated the presence of an allele specific β-catenin activity level conferred by these mutational variants, which cannot be explained by the current model of β-catenin regulation. The results of this study imply a fundamental difference between these mutations, the existence of a genotype-phenotype correlation based on β-catenin activity, and challenges the prevailing dogma of β-catenin regulation, thus emphasizing the need to further examine the underlying mechanistic process involved in the observed differential phenotypic response.

http://hdl.handle.net/1842/35817

en

The University of Edinburgh

β-catenin activity

β-catenin mutations

CRISPR/Cas9

Genotype-phenotype correlations in β-catenin

Thesis or Dissertation

Doctoral

PhD Doctor of Philosophy