With the completion of sequencing of the human and mouse genomes, the comprehensive functional annotation and analysis of mammalian genes have become achievable goals. Among the multitude of experimental approaches addressing gene function, the most relevant for extrapolation to human genetic disease is the generation and analysis of mutation-induced phenotypes. Several model organisms have been used in mutagenesis approaches, but the mouse offers particular advantages because its genome structure and organization is closely related to the human genome. Most importantly, the availability of mouse embryonic stem cells (ESCs), which grow indefinitely in tissue culture, allows the generation of ESC lines with defined mutations that can be used for production of mutant mice.
Within the framework of the International Mouse Knock Out Consortium (IKMC), my laboratory participates in the systematic knock-out of mouse genes in ESCs using high throughput sequence-based approaches, such as gene trapping and gene targeting. The overall goal of the effort is to assemble a library of ES cell lines harboring conditional mutations in every single gene of the mouse genome, and to make these cell lines available to the scientific community for production of mutant mice.
Knockouts primarily address the function of single genes. However, since altered pathways rather than single genes are more often responsible for disease, understanding the role of these genes in relevant pathways is essential. This is most effectively achieved using tag-based assays for systematic protein localization and protein-protein interaction studies. Ideally, protein tags are introduced into the genomic locus of the gene of interest to ensure expression from endogenous control elements. Within the framework of the DiGtoP consortium (From Disease Genes to Proteins), my laboratory is developing tools for in situ tagging of disease related proteins in mouse and human ESCs. The protein tagged ESC lines will enable systematic protein localization and protein-protein interaction studies under physiological conditions and will be useful for applications ranging from comparative mouse/human proteome analysis in ESC differentiation cultures to the definition of tissue-specific proteomes in mice.
A further interest of the laboratory is in the biology of sestrins and their significance for human disease. Mammalian cells express three isoforms (sesn1-3) whose functions are still poorly understood. Besides being antioxidant proteins, sestrins control several intracellular signal transduction pathways including the TGF-β and mTOR pathways. Based on our observation that the mutational inactivation of sesn2 in a transgenic mouse model of COPD partially rescues the emphysema phenotype by inducing the expression deposition of elastin in the lung, the laboratory is committed to resolve the responsible molecular mechanisms. Moreover, in collaboration with the University of Gießen Lung Center, we are exploring the possibility of sesn2 as a novel target in the treatment of COPD.
Finally, in collaboration with the Department of Hematology, we are analyzing the role of sestrins in the regulation of receptor tyrosine kinases and their oncogenic variants in hematopoietic cells, with the ultimate goal to identify novel targets for cancer therapy.
The Laboratory is part of the National German Genome Network (NGFNplus) and the European Conditional Mouse Mutagenesis (EUCOMM) program. It receives funding from the Bundesministerium für Bildung und Forschung (BMBF), the European Union, the Deutsche Forschungsgemeinschaft and the Boehringer Ingelheim Stiftung.
