Ubiquitinylation of α-synuclein by carboxyl terminus Hsp70-interacting protein (CHIP) is regulated by Bcl-2-associated athanogene 5 (BAG5).
Ubiquitinylation of α-synuclein by carboxyl terminus Hsp70-interacting protein (CHIP) is regulated by Bcl-2-associated athanogene 5 (BAG5).
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DOI：
10.1371/journal.pone.0014695
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发表时间：
2011-02-16
期刊：
影响因子：
3.7
通讯作者：
McLean PJ
中科院分区：
文献类型：
--
作者：
Kalia LV;Kalia SK;Chau H;Lozano AM;Hyman BT;McLean PJ
帕金森病(PD)是一种常见的神经退行性疾病，在这种情况下，蛋白质稳态或蛋白质稳态的异常可能导致蛋白质α-突触核蛋白(α-SYN)的积聚。编码α-SYN基因的突变或倍增被认为是导致遗传形式的帕金森病的原因，该基因的多态是最近确定的特发性帕金森病的危险因素。α-SYN是路易小体的主要成分，路易小体是细胞内的蛋白质包涵体，是大多数形式帕金森病的病理特征。最近的证据表明，α-SYN可以自我结合成可溶性寡聚体，并与这些α-SYN寡聚体参与细胞死亡有关。我们先前已经证明，热休克蛋白70相互作用蛋白的羧基末端(CHIP)是一种具有E3泛素连接酶活性的辅助伴侣分子，可能会减少有毒的α-SYN寡聚体的水平。在这里，我们证明了α-SYN在体外和细胞内都是由CHIP泛素化的。我们发现，芯片泛素化的产物包括单泛素化和多泛素化的α-SYN。我们还证明了CHIP和α-SYN存在于大脑中与辅助伴侣BCL-2相关的基因5(BAG5)的蛋白质复合体中。CHIP与BAG5的相互作用是由Hsp70介导的，Hsp70与CHIP的四肽重复结构域和BAG5的袋区结合。热休克蛋白70介导的BAG5与CHIP的结合导致CHIP E3泛素连接酶活性的抑制，从而降低α-SYN泛素化。此外，我们使用基于荧光素酶的α-SYN寡聚的蛋白质片段互补分析来研究CHIP对活细胞中α-SYN寡聚体的调控。我们证明，BAG5减弱了CHIP抑制α-SYN寡聚的能力，而非泛素化的α-SYN具有更强的寡聚倾向。因此，我们的结果确定CHIP是α-SYN的E3泛素连接酶，并提示BAG5作为CHIP E3泛素连接酶活性的调节器具有新的功能，这可能与CHIP介导的α-SYN寡聚调控有关。
Parkinson's disease (PD) is a common neurodegenerative condition in which abnormalities in protein homeostasis, or proteostasis, may lead to accumulation of the protein α-synuclein (α-syn). Mutations within or multiplications of the gene encoding α-syn are known to cause genetic forms of PD and polymorphisms in the gene are recently established risk factors for idiopathic PD. α-syn is a major component of Lewy bodies, the intracellular proteinaceous inclusions which are pathological hallmarks of most forms of PD. Recent evidence demonstrates that α-syn can self associate into soluble oligomeric species and implicates these α-syn oligomers in cell death. We have previously shown that carboxyl terminus of Hsp70-interacting protein (CHIP), a co-chaperone molecule with E3 ubiquitin ligase activity, may reduce the levels of toxic α-syn oligomers. Here we demonstrate that α-syn is ubiquitinylated by CHIP both in vitro and in cells. We find that the products from ubiquitinylation by CHIP include both monoubiquitinylated and polyubiquitinylated forms of α-syn. We also demonstrate that CHIP and α-syn exist within a protein complex with the co-chaperone bcl-2-associated athanogene 5 (BAG5) in brain. The interaction of CHIP with BAG5 is mediated by Hsp70 which binds to the tetratricopeptide repeat domain of CHIP and the BAG domains of BAG5. The Hsp70-mediated association of BAG5 with CHIP results in inhibition of CHIP E3 ubiquitin ligase activity and subsequently reduces α-syn ubiquitinylation. Furthermore, we use a luciferase-based protein-fragment complementation assay of α-syn oligomerization to investigate regulation of α-syn oligomers by CHIP in living cells. We demonstrate that BAG5 mitigates the ability of CHIP to reduce α-syn oligomerization and that non-ubiquitinylated α-syn has an increased propensity for oligomerization. Thus, our results identify CHIP as an E3 ubiquitin ligase of α-syn and suggest a novel function for BAG5 as a modulator of CHIP E3 ubiquitin ligase activity with implications for CHIP-mediated regulation of α-syn oligomerization.