The Doa4 deubiquitinating enzyme is required for ubiquitin homeostasis in yeast
The Doa4 deubiquitinating enzyme is required for ubiquitin homeostasis in yeast
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DOI：
10.1091/mbc.10.8.2583
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发表时间：
1999-08-01
影响因子：
3.3
通讯作者：
Hochstrasser, M
中科院分区：
文献类型：
--
作者：
Swaminathan, S;Amerik, AY;Hochstrasser, M
泛素与细胞蛋白的结合通常以26S蛋白酶体为目标进行降解。此外，细胞表面蛋白的泛素化刺激它们的内吞作用，并最终在液泡或溶酶体中降解。在酿酒酵母中，泛素是一种长寿的蛋白质，因此它必须从与之相连的蛋白质分解中间体中有效地回收利用。我们以前发现了一种酵母去泛素酶Doa4，它在蛋白酶体依赖于泛素的蛋白分解过程中起着核心作用。生化和遗传学数据表明，Doa4的作用与蛋白酶体的作用密切相关。在这里，我们提供的证据表明，Doa4是从泛素化底物中回收泛素所必需的，泛素底物针对蛋白酶体，令人惊讶的是，也针对液泡。在doa4 Delta突变体中，泛素在某些条件下被强烈耗尽，最明显的是当细胞接近静止时。在静止期的DOA4Delta细胞中，泛素的耗尽先于细胞活力的显著丧失。Doa4 Delta细胞的这种活性丧失和其他几个缺陷可以通过提供额外的泛素来挽救。泛素在突变体中变得枯竭，因为它的降解速度比野生型细胞快得多。蛋白酶体的突变或空泡蛋白分解或内吞作用的失活可以部分抑制泛素的异常降解。我们认为Doa4有助于从蛋白酶体结合的泛素化中间体和膜蛋白中回收泛素，这些蛋白将在液泡中被破坏。
Attachment of ubiquitin to cellular proteins frequently targets them to the 26S proteasome for degradation. in addition, ubiquitination of cell surface proteins stimulates their endocytosis and eventual degradation in the vacuole or lysosome. In the yeast Saccharomyces cerevisiae, ubiquitin is a long-lived protein, so it must be efficiently recycled from the proteolytic intermediates to which it becomes linked. We identified previously a yeast deubiquitinating enzyme, Doa4, that plays a central role in ubiquitin-dependent proteolysis by the proteasome. Biochemical and genetic data suggest that Doa4 action is closely linked to that of the proteasome. Here we provide evidence that Doa4 is required for recycling ubiquitin from ubiquitinated substrates targeted to the proteasome and, surprisingly, to the vacuole as well. In the doa4 Delta mutant, ubiquitin is strongly depleted under certain conditions, most notably as cells approach stationary phase. Ubiquitin depletion precedes a striking loss of cell viability in stationary phase doa4 Delta cells. This loss of viability and several other defects of doa4 Delta cells are rescued by provision of additional ubiquitin. Ubiquitin becomes depleted in the mutant because it is degraded much more rapidly than in wild-type cells. Aberrant ubiquitin degradation can be partially suppressed by mutation of the proteasome or by inactivation of vacuolar proteolysis or endocytosis. We propose that Doa4 helps recycle ubiquitin from both proteasome-bound ubiquitinated intermediates and membrane proteins destined for destruction in the vacuole.