Laboratory of Protein Metabolism, Tokyo Metropolitan Institute of Medical Science, Setagaya, Tokyo 156-8506, Japan.
Laboratory of Protein Metabolism, Tokyo Metropolitan Institute of Medical Science, Setagaya, Tokyo 156-8506, Japan.
Mol Cell. 2017 May 18;66(4):488-502.e7. doi: 10.1016/j.molcel.2017.04.024.
Ubiquitin-binding domain (UBD) proteins regulate numerous cellular processes, but their specificities toward ubiquitin chain types in cells remain obscure. Here, we perform a quantitative proteomic analysis of ubiquitin linkage-type selectivity of 14 UBD proteins and the proteasome in yeast. We find that K48-linked chains are directed to proteasomal degradation through selectivity of the Cdc48 cofactor Npl4. Mutating Cdc48 results in decreased selectivity, and lacking Rad23/Dsk2 abolishes interactions between ubiquitylated substrates and the proteasome. Among them, only Npl4 has K48 chain specificity in vitro. Thus, the Cdc48 complex functions as a K48 linkage-specifying factor upstream of Rad23/Dsk2 for proteasomal degradation. On the other hand, K63 chains are utilized in endocytic pathways, whereas both K48 and K63 chains are found in the MVB and autophagic pathways. Collectively, our results provide an overall picture of the ubiquitin network via UBD proteins and identify the Cdc48-Rad23/Dsk2 axis as a major route to the proteasome.
泛素结合结构域（UBD）蛋白调节许多细胞过程，但它们在细胞中对不同类型泛素链的特异性仍然不清楚。在这里，我们对酵母中 14 种 UBD 蛋白和蛋白酶体的泛素连接类型选择性进行了定量蛋白质组学分析。我们发现，Cdc48 辅助因子 Npl4 对 K48 连接链具有选择性，使其定向到蛋白酶体降解。Cdc48 突变会降低选择性，而缺乏 Rad23/Dsk2 会破坏泛素化底物与蛋白酶体之间的相互作用。其中，只有 Npl4 在体外具有 K48 链特异性。因此，Cdc48 复合物在 Rad23/Dsk2 上游作为 K48 连接指定因子，参与蛋白酶体降解。另一方面，K63 链用于内吞途径，而 K48 和 K63 链都存在于 MVB 和自噬途径中。总之，我们的研究结果提供了一个通过 UBD 蛋白的泛素网络的整体画面，并确定了 Cdc48-Rad23/Dsk2 轴是蛋白酶体的主要途径。
