A novel in vivo overexpression screen for cell cycle and growth regulating genes in Drosophila
Final Report Abstract
To identify genes that regulate cell growth, cell cycle progression and proliferation in vivo, I created a library of transgenic strains in the fruit fly Drosophila melanogaster. Each strain contains one of 650 open reading frames (ORFs) inserted into a specific locus of the Drosophila genome. We developed a versatile expression system that facilitates in vivo exchange of promoter- and tagsequences. The ORFs that were used were identified by a RNAi based loss-offunction (lf) approach in Drosophila S2 cells to function in cell cycle and cell size regulation as well as a variety of related mechanisms. I screened the resulting 650 fly lines for a variety of growth related phenotypes. This work provided a comprehensive and systematic in vivo analysis of growth relevant genes in Drosophila. Furthermore, by bioinformatic integration of the independent screening results I could successfully apply a meta-analysis of the data, which allowed me to identify high confidence candidates for novel components of the Wnt/Wg-signaling pathway. Validation of these candidates led to the identification of three kinases that function as novel pathway components. Strikingly, these kinases lack a wg-phenotype upon RNAi mediated knock-down, likely due to genetic redundancy. Thus, these genes would have been missed in conventional loss-of-function screens, but could only be identified by our misexpression approach. Mutations within the human homologs of the three kinases are associated with different types of cancer and our work suggests a misregulation of the Wnt-pathway as a cause for these malignancies.
