TFTag: A novel library of tagged transcription factors in Drosophila

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Every multicellular organism consists of dozens or even hundreds of different cell types that are defined by their different protein repertoires. The building plans for the proteins are encoded in the form of genes in the DNA, as are the instructions of where and when to make them. This process, referred to as gene expression, is highly dynamic and is constantly adjusted during development or in response to, e.g., environmental changes. Each cell type has its characteristic pattern of gene expression, and mis-regulation is associated with various diseases, including cancer. Therefore, gene expression must be tightly regulated to ensure that each cell has the correct protein content. This is largely done by a specific class of proteins, the transcription factors (TFs), which can bind to DNA and cooperate with each other to influence which genes are transcribed in what quantities. This in turn determines which proteins are present in which cells and in which amount. TFs are, accordingly, studied by a wide range of scientists in the life and medical sciences.Research on TFs mainly seeks to uncover information about their expression level, localisation, binding sites in the genome, interaction partners, and effects of a loss of function. Most of these analyses rely heavily on the use of antibodies, the production of which is costly, difficult, and ethically questionable. Moreover, methods involving antibodies are geared towards fixed biological materials, which makes the recapitulation of dynamic processes challenging. A common solution is the use of genetic tricks to express a version of the TF that is fused to another protein, which can, in turn, be readily detected and is, accordingly, called a tag. However, tags are often suitable for only a single or few methods or otherwise are large and might interfere with the function of the TF. Moreover, the additional expression of the tagged TF can induce artefacts. Also, tools for the expression of such tagged constructs are only available for a subset of TFs.The fruit fly, Drosophila melanogaster, has long served as one of the prime model organisms to study basic biological processes. It benefits from a large, well-connected worldwide community of researchers and the availability of vast genetic and molecular resources. Importantly, it allows investigations in vivo, and the creation of transgenic lines for the generation of novel tools is straightforward.Here we propose to create a biological and bioinformatic resource consisting of a library of transgenic fly lines in which all Drosophila TF genes have been tagged at their native genomic locus. We will also establish a database with basic data on the expression and DNA binding of a subset of TFs that have not previously been studied. Our resources will alleviate many of the shortcomings of previous libraries of tagged proteins: 1) Our tagging strategy allows for the easy exchange of the tag, making each TF accessible for various downstream applications; 2) We will create individual transgenic lines in which either end of a TF has been tagged, which reduces the risk of an interference of the tag with protein function; 3) For TFs that are expressed with alternative ends, we will create a line for each end, allowing the study of different versions of each TF. We will distribute the fly lines free of charge to members of the research community, and the database will, likewise, be free to use. These resources will open the possibility to study any Drosophila TF with the methods of choice in any biological process imaginable and allow scientists to easily identify potential candidates among the many unstudied TFs. We expect that our resource will greatly advance research in the field of gene regulation.