Department for Bioinformatics and Functional Genomics, Division of Theoretical Bioinformatics, German Cancer Research Center (DKFZ), Institute for Pharmacy and Molecular Biotechnology (IPMB) and BioQuant, Heidelberg, Germany.
Cell Death Differ. 2013 Apr;20(4):599-610. doi: 10.1038/cdd.2012.156. Epub 2013 Jan 11.
Apoptosis occurs through a tightly regulated cascade of caspase activation. In the context of extrinsic apoptosis, caspase-8 is activated by dimerization inside a death receptor complex, cleaved by auto-proteolysis and subsequently released into the cytosol. This fully processed form of caspase-8 is thought to cleave its substrates BID and caspase-3. To test if the release is required for substrate cleavage, we developed a novel approach based on localization probes to quantitatively characterize the spatial-temporal activity of caspases in living single cells. Our study reveals that caspase-8 is significantly more active at the plasma membrane than within the cytosol upon CD95 activation. This differential activity is controlled by the cleavage of caspase-8 prodomain. As a consequence, targeting of caspase-8 substrates to the plasma membrane can significantly accelerate cell death. Subcellular compartmentalization of caspase-8 activity may serve to restrict enzymatic activity before mitochondrial pathway activation and offers new possibilities to interfere with apoptotic sensitivity of the cells.
细胞凋亡是通过 caspase 激活的严格调控级联反应发生的。在外源性细胞凋亡中，caspase-8 通过死亡受体复合物内的二聚化而被激活，通过自身蛋白水解切割，随后被释放到细胞质中。这种完全加工形式的 caspase-8 被认为可以切割其底物 BID 和 caspase-3。为了测试释放是否是底物切割所必需的，我们开发了一种基于定位探针的新方法，用于定量表征活单细胞中 caspase 的时空活性。我们的研究表明，在 CD95 激活后，caspase-8 在质膜上的活性明显高于细胞质。这种差异活性受 caspase-8 前导肽的切割控制。因此，将 caspase-8 底物靶向质膜可以显著加速细胞死亡。caspase-8 活性的亚细胞区室化可能有助于在线粒体途径激活之前限制酶的活性，并为干预细胞凋亡敏感性提供了新的可能性。
