Su Chen-Hsien, McStay Gavin P, Tzagoloff Alexander
Department of Biological Sciences, Columbia University, New York, NY 10027.
Mol Biol Cell. 2014 Apr;25(7):965-76. doi: 10.1091/mbc.E13-10-0575. Epub 2014 Jan 29.
Yeast cytochrome oxidase (COX) was previously inferred to assemble from three modules, each containing one of the three mitochondrially encoded subunits and a different subset of the eight nuclear gene products that make up this respiratory complex. Pull-down assays of pulse-labeled mitochondria enabled us to characterize Cox3p subassemblies that behave as COX precursors and contain Cox4p, Cox7p, and Cox13p. Surprisingly, Cox4p is a constituent of two other complexes, one of which was previously proposed to be an intermediate of Cox1p biogenesis. This suggests that Cox4p, which contacts Cox1p and Cox3p in the holoenzyme, can be incorporated into COX by two alternative pathways. In addition to subunits of COX, some Cox3p intermediates contain Rcf1p, a protein associated with the supercomplex that stabilizes the interaction of COX with the bc1 (ubiquinol-cytochrome c reductase) complex. Finally, our results indicate that although assembly of the Cox1p module is not contingent on the presence of Cox3p, the converse is not true, as none of the Cox3p subassemblies were detected in a mutant blocked in translation of Cox1p. These studies support our proposal that Cox3p and Cox1p are separate assembly modules with unique compositions of ancillary factors and subunits derived from the nuclear genome.
酵母细胞色素氧化酶（COX）先前被推断由三个模块组装而成，每个模块包含三个线粒体编码亚基中的一个以及构成该呼吸复合体的八个核基因产物的不同子集。对脉冲标记线粒体的下拉分析使我们能够表征作为COX前体且包含Cox4p、Cox7p和Cox13p的Cox3p亚组装体。令人惊讶的是，Cox4p是另外两个复合体的组成部分，其中一个先前被认为是Cox1p生物合成的中间体。这表明在全酶中与Cox1p和Cox3p接触的Cox4p可以通过两种替代途径整合到COX中。除了COX的亚基外，一些Cox3p中间体还包含Rcf1p，一种与超复合体相关的蛋白质，它稳定了COX与bc1（泛醇 - 细胞色素c还原酶）复合体的相互作用。最后，我们的结果表明，虽然Cox1p模块的组装不依赖于Cox3p的存在，但反之则不然，因为在Cox1p翻译受阻的突变体中未检测到任何Cox3p亚组装体。这些研究支持了我们的提议，即Cox3p和Cox1p是具有独特辅助因子和源自核基因组的亚基组成的独立组装模块。
