Dissection of molecular Mechanisms regulating Pexophagy in human Cells
Final Report Abstract
This project was initially designed to shed light upon the genes and mechanisms regulating autophagy-mediated degradation of peroxisomes (a process termed ‘pexophagy’) in human cells. The conceptual and methodological difficulties that we encountered during an initial establishment and validation phase prompted us to change the general direction of the project slightly: We proceeded by focusing on several aspects of general and selective autophagy in yeast and mammals, and incorporated a systems biology based gene network approach to assess autophagy on a global level. The salient findings from this study emphasize that a combination of network analysis strategies and assessment of evolutionary conservation of the associated genes and processes represents a powerful toolset for experiment design, interpretation of large data sets and hypothesis generation. In short, we performed a meta-analysis of functional data and established a gene interaction network comprising over 1200 human autophagy related genes and several thousands of interactions of various types. The resulting network could be utilized for interpretation of phospho-proteomic data in the context of Nemo-dependent signaling, for analysis of gene expression during bacterial cytoinvasion, and to understand the crosstalk between autophagy and cell adhesion. Importantly, network characteristics were used to identify promising candidates for a focused siRNA screen for pexophagy and bulk autophagy using high content imaging. Results from this screen and similar approaches formed the basis for functional follow up studies both in yeast and human cells, and have contributed to the characterization of a novel pexophagy specific modulator gene that is conserved from yeast to man and may represent a molecular target for treatment of peroxisome biogenesis disorders or neurodegenerative diseases. Surprisingly, we found that, in this context, simple unicellular organisms like yeast can effectively be used to unravel complex processes that are of pathophysiological relevance for higher eukaryotes. In other words, following up on predictions from yeast-based models can help us understand the molecular mechanisms underlying human diseases. The entire autophagy gene system has not been characterized to a comparable level of detail before, and our data set (which comprises information about the respective gene orthologs in almost 100 model organisms) represents a versatile tool for refining research strategies in different fields. Furthermore, our strategy facilitated the expansion of our research from pexophagy to several aspects of selective and bulk autophagy, thus enabling us to contribute to the characterization of novel risk factors for Crohn disease, to unravel mechanisms underlying neuropathologies, and to decipher aspects of crosstalk between autophagy and immune signaling. In summary, we used a systems biology approach in combination with classical yeast and mammalian cell biology to describe aspects of selective autophagy pathways with various links to human disorders (neurodegeneration, chronic inflammation) that might result in patent filing efforts and/or increased public interest in the field of selective autophagy.
Publications
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(2010) A balancing act for autophagin. J Clin Invest 120, 2273-2276
Till, A., and Subramani, S.
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(2011) The complex interplay of NOD-like receptors and the autophagy machinery in the pathophysiology of Crohn disease. Eur J Cell Biol 90, 593-602
Billmann-Born, S., Lipinski, S., Bock, J., Till, A., Rosenstiel, P., and Schreiber, S.
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(2012) Extracellular cathepsin K exerts antimicrobial activity and is protective against chronic intestinal inflammation in mice. Gut. 2012 Mar 22
Sina, C., Lipinski, S., Gavrilova, O., Aden, K., Rehman, A., Till, A., Rittger, A., Podschun, R., Meyer- Hoffert, U., Haesler, R., Midtling, E., Putsep, K., McGuckin, M. A., Schreiber, S., Saftig, P., and Rosenstiel, P.
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(2012) Pexophagy: the selective degradation of peroxisomes. Int J Cell Biol 2012, 512721
Till, A., Lakhani, R., Burnett, S. F., and Subramani, S.
