Zhou Liying, Chang Donald C
Department of Biology, the Hong Kong University of Science and Technology, Clear Water Bay, Kowloon, Hong Kong.
J Cell Sci. 2008 Jul 1;121(Pt 13):2186-96. doi: 10.1242/jcs.024703. Epub 2008 Jun 10.
Bax and Bak are known to play a central role in facilitating the release of mitochondrial intermembrane proteins during apoptosis. The detailed mechanism, however, is still not clear. Using live cell imaging techniques, we showed here that Bax underwent four distinct stages of dynamic redistribution during UV-induced apoptosis. At stage I, Bax was distributed diffusely in the cytosol. About an hour after UV treatment at stage II, Bax started to translocate to mitochondria and distributed uniformly at the mitochondrial outer membrane (MOM). Within a few minutes, at stage III, Bax and Bak began to form small complexes at the MOM. Later, at stage IV, these Bax and Bak complexes expanded to become large clusters. We found that the formation of Bax-Bak small complexes at stage III was responsible for permeabilizing the MOM to release cytochrome c and Smac. Using a FRET technique, we further showed that Bax binds to Bak within the complex formed at the MOM during stage III. Finally, using a quantitative fluorescence measurement, we determined that the Bax-Bak complex was about 0.25 microm wide and composed of more than 100 protein molecules. These findings suggest that the Bax-Bak structure responsible for releasing mitochondrial proteins during apoptosis is not channel-like.
已知Bax和Bak在细胞凋亡过程中促进线粒体膜间蛋白释放方面发挥核心作用。然而，具体机制仍不清楚。我们利用活细胞成像技术在此表明，在紫外线诱导的细胞凋亡过程中，Bax经历了四个不同阶段的动态重新分布。在第一阶段，Bax分散分布于细胞质中。在第二阶段紫外线处理约一小时后，Bax开始转位至线粒体，并均匀分布于线粒体外膜（MOM）。几分钟内，在第三阶段，Bax和Bak开始在MOM处形成小复合物。随后，在第四阶段，这些Bax和Bak复合物扩展成为大簇。我们发现第三阶段Bax-Bak小复合物的形成导致MOM通透，从而释放细胞色素c和Smac。利用荧光共振能量转移（FRET）技术，我们进一步表明在第三阶段形成于MOM的复合物中Bax与Bak结合。最后，通过定量荧光测量，我们确定Bax-Bak复合物宽约0.25微米，由100多个蛋白质分子组成。这些发现表明，在细胞凋亡过程中负责释放线粒体蛋白的Bax-Bak结构并非通道样结构。
