A bacterial type III secretion-based protein delivery tool for broad applications in cell biology
| Autor: | Marlise Amstutz, Markus Affolter, Simon Ittig, Shyan Huey Low, Loïc Sauteur, Cécile Arrieumerlou, Erich A. Nigg, Christoph Kasper, M. Alessandra Vigano, Alexander Schmidt, Christoph Schmutz, Guy R. Cornelis |
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| Rok vydání: | 2015 |
| Předmět: |
Proteases
Cell Membrane Permeability Proteome Truncated BID Recombinant Fusion Proteins Molecular Sequence Data Apoptosis Tools Mice Drug Delivery Systems Ubiquitin Type III Secretion Systems Animals Humans Secretion Phosphorylation Research Articles Zebrafish biology Phosphoproteomics 3T3 Cells Cell Biology Fusion protein Cell biology Transport protein Protein Transport biology.protein Apoptosis Regulatory Proteins Protein Processing Post-Translational Bacterial Outer Membrane Proteins HeLa Cells |
| Zdroj: | The Journal of Cell Biology |
| ISSN: | 1540-8140 0021-9525 |
| Popis: | Protein delivery based on bacterial type III secretion allows controllable injection of bacterial, viral, and human proteins into target cells, functional interaction studies, targeting of proteins to subcellular locations, and systems-level signaling analysis of a given translocated protein. Methods enabling the delivery of proteins into eukaryotic cells are essential to address protein functions. Here we propose broad applications to cell biology for a protein delivery tool based on bacterial type III secretion (T3S). We show that bacterial, viral, and human proteins, fused to the N-terminal fragment of the Yersinia enterocolitica T3S substrate YopE, are effectively delivered into target cells in a fast and controllable manner via the injectisome of extracellular bacteria. This method enables functional interaction studies by the simultaneous injection of multiple proteins and allows the targeting of proteins to different subcellular locations by use of nanobody-fusion proteins. After delivery, proteins can be freed from the YopE fragment by a T3S-translocated viral protease or fusion to ubiquitin and cleavage by endogenous ubiquitin proteases. Finally, we show that this delivery tool is suitable to inject proteins in living animals and combine it with phosphoproteomics to characterize the systems-level impact of proapoptotic human truncated BID on the cellular network. |
| Databáze: | OpenAIRE |
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