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Molecular regulation of caspase activation - role of interacting proteins


Research objectives and content Apoptotic cell death is a general phenomenon occurring in all multicellular organisms. It has been described as a form of cellular suicide since death appears to result from the induction of active processes within the cell. Morphological features of apoptosis are very distinct. Induction of these morphological changes is associated with increased activity of lipases, nucleases and different classes of proteases. The resultant macromolecular degradation facilitates neat packaging of intracellular contents into apoptotic bodies thus protecting neighbouring cells from damage. The caspase family of aspartic acid-specific cysteine proteases, which are activated during apoptosis in numerous cell systems, cleave several proteins localised to the cytosol, cytoskeleton and nucleus. There are at least 10 members of this family, sub-divided according to their substrate specificity and function. Caspases are normally present in cells as pro-enzymes and are activated by cleavage at specific sites. Caspase-3, which is activated in almost all cases of apoptosis reported to date, is thought to be one of the main regulators of apoptosis. The proposed project is based on our recent findings that pro-caspase-3 is localised not only to cytosol but also to mitochondria. Our findings are interesting in view of the central role played by the mitochondria in the induction of apoptosis and they raise a number of questions regarding the biological significance of mitochondrial pro-caspase-3. These include, the mechanism of transport of pro-caspase-3 into the mitochondria, possible mechanisms of activation of mitochondrial caspase-3 and the mechanisms of active caspase-3 translocation to the nuclei where it exerts its proteolytic activity on a number of substrates. These questions will be addressed during the course of this project. Training content (objective, benefit and expected impact) I have been at the host institute for the past eight months and have found the atmosphere very stimulating and collaborative. I chose this group because of its excellent track record, its experience in the field, inter disciplinary research interests and the facilities available. The fact that this group puts a great deal of emphasis on training and familiarisation with new techniques and equipment has been more appealing to me than I previously thought. I have already learnt a great deal about the enzymes of interest in apoptosis and specific enzyme assays. Furthermore, I have also gained experience in protein purification using FPLC, high molecular weight DNA fragmentation, and various microscopy techniques. For the proposed project I will learn 2-dimensional gel electrophoresis, in vitro translation/transcription, protein sequencing, and development of monoclonal antibodies. Overall the planned research, which is estimated to take 24 months, will not only give me a deeper knowledge and a wider competence in the areas mentioned but also give me more experience in interacting and collaborating with other researchers. Links with industry / industrial relevance (22)

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Aristotle University of Thessaloniki
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Analytical Chemistry Lab
54006 Thessaloniki

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