PAM forms an atypical SCF ubiquitin ligase complex that ubiquitinates and degrades NMNAT2
PAM forms an atypical SCF ubiquitin ligase complex that ubiquitinates and degrades NMNAT2
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DOI：
10.1074/jbc.ra118.002176
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发表时间：
2018-09-07
影响因子：
4.8
通讯作者：
Grill, Brock
中科院分区：
文献类型：
--
作者：
Desbois, Muriel;Crawley, Oliver;Grill, Brock
PHR (PAM/Highwire/RPM-1) 蛋白是保守的 RING E3 泛素连接酶，在轴突终止和突触形成以及轴突变性等发育过程中发挥作用。目前，我们对 PHR 蛋白如何形成泛素连接酶复合物的理解仍然不完整。尽管遗传学研究表明 NMNAT2 是 PHR 蛋白在轴突变性中发挥功能的重要介质，但 PHR 蛋白如何抑制 NMNAT2 仍不清楚。在这里，我们破译了人类 PHR 蛋白 PAM（也称为 MYCBP2）如何形成非规范的 Skp/Cullin/F-box (SCF) 复合物的生化基础，该复合物包含 F-box 蛋白 FBXO45 和 SKP1 但缺乏 CUL1。我们证明 FBXO45 不仅在底物识别中发挥作用，而且对于 PAM/FBXO45/SKP1 复合物的组装也很重要。有趣的是，我们证明了 SKP1 作为目标识别模块的辅助组件的新作用，可增强 FBXO45 与 NMNAT2 的结合。最后，我们提供了生化证据表明 PAM 多泛素化 NMNAT2 并通过蛋白酶体调节 NMNAT2 蛋白稳定性和降解。
PHR (PAM/Highwire/RPM-1) proteins are conserved RING E3 ubiquitin ligases that function in developmental processes, such as axon termination and synapse formation, as well as axon degeneration. At present, our understanding of how PHR proteins form ubiquitin ligase complexes remains incomplete. Although genetic studies indicate NMNAT2 is an important mediator of PHR protein function in axon degeneration, it remains unknown how PHR proteins inhibit NMNAT2. Here, we decipher the biochemical basis for how the human PHR protein PAM, also called MYCBP2, forms a noncanonical Skp/Cullin/F-box (SCF) complex that contains the F-box protein FBXO45 and SKP1 but lacks CUL1. We show FBXO45 does not simply function in substrate recognition but is important for assembly of the PAM/FBXO45/SKP1 complex. Interestingly, we demonstrate a novel role for SKP1 as an auxiliary component of the target recognition module that enhances binding of FBXO45 to NMNAT2. Finally, we provide biochemical evidence that PAM polyubiquitinates NMNAT2 and regulates NMNAT2 protein stability and degradation by the proteasome.