Dorstyn Loretta, Mills Kathryn, Lazebnik Yuri, Kumar Sharad
Hanson Institute, Adelaide, Australia 5000.
J Cell Biol. 2004 Nov 8;167(3):405-10. doi: 10.1083/jcb.200408054.
In Drosophila, activation of the apical caspase DRONC requires the apoptotic protease-activating factor homologue, DARK. However, unlike caspase activation in mammals, DRONC activation is not accompanied by the release of cytochrome c from mitochondria. Drosophila encodes two cytochrome c proteins, Cytc-p (DC4) the predominantly expressed species, and Cytc-d (DC3), which is implicated in caspase activation during spermatogenesis. Here, we report that silencing expression of either or both DC3 and DC4 had no effect on apoptosis or activation of DRONC and DRICE in Drosophila cells. We find that loss of function mutations in dc3 and dc4, do not affect caspase activation during Drosophila development and that ectopic expression of DC3 or DC4 in Drosophila cells does not induce caspase activation. In cell-free studies, recombinant DC3 or DC4 failed to activate caspases in Drosophila cell lysates, but remarkably induced caspase activation in extracts from human cells. Overall, our results argue that DARK-mediated DRONC activation occurs independently of cytochrome c.
在果蝇中，顶端半胱天冬酶DRONC的激活需要凋亡蛋白酶激活因子同源物DARK。然而，与哺乳动物中的半胱天冬酶激活不同，DRONC的激活并不伴随着细胞色素c从线粒体的释放。果蝇编码两种细胞色素c蛋白，主要表达的Cytc-p（DC4）和与精子发生过程中的半胱天冬酶激活有关的Cytc-d（DC3）。在此，我们报道，沉默DC3和DC4其中之一或二者的表达，对果蝇细胞中的凋亡或DRONC和DRICE的激活均无影响。我们发现，dc3和dc4的功能缺失突变不影响果蝇发育过程中的半胱天冬酶激活，并且在果蝇细胞中异位表达DC3或DC4不会诱导半胱天冬酶激活。在无细胞研究中，重组DC3或DC4未能在果蝇细胞裂解物中激活半胱天冬酶，但在人细胞提取物中却能显著诱导半胱天冬酶激活。总体而言，我们的结果表明，DARK介导的DRONC激活独立于细胞色素c发生。
