Gene regulation is largely controlled at the transcriptional level, where transcription factors (TFs) interact with regulatory elements in the DNA to repress or enhance activity of RNA polymerase II. The study of TFs involves analysis of expression patterns, genomic binding sites, and interaction partners, all of which rely on the use of specific antibodies. However, the production of antibodies is expensive, time-consuming, and ethically questionable. In Drosophila melanogaster, one of the prime model organisms for the analysis of basic biological processes, a common solution is the use of tagged versions of TFs. Tags encode small peptides, fluorescent proteins, or enzymatic domains and allow for various downstream applications. Fly strain libraries allowing for analysis of tagged proteins have been created in recent years but they only cover a small and largely already well-studied subset of the 753 known or predicted Drosophila TFs. Here, we propose to create a biological and bioinformatic resource to greatly advance the analysis of TFs in Drosophila and beyond. This resource will consist of three parts: 1) a library of plasmids for the CRISPR-mediated tagging of all Drosophila TFs; 2) a collection of fly lines in which the tags have been introduced at endogenous loci; 3) a database with basic information about the plasmids and tagged lines, as well as expression and binding data for a subset of previously unstudied TFs. We plan to tag TFs at every annotated N and C-terminus. This will alleviate problems that might arise from interference of the tag with one terminus and at the same time allow analysis of all TF variants with different termini. Moreover, the primary sfGFP will be easily exchangeable by crossing to a donor line for any other tag that might be desired. This will allow any downstream analysis imaginable and substantially broaden the user base. Furthermore, our database will allow researchers to easily select candidate TFs to study in more detail.
TFTag: A novel library of tagged transcription factors in Drosophila
Dr Korneel Hens
Oxford Brookes University