Landscape of the PARKIN-dependent ubiquitylome in response to mitochondrial depolarization.
Landscape of the PARKIN-dependent ubiquitylome in response to mitochondrial depolarization.
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作者：
帕金（PARK2）泛素连接酶及其调节激酶PINK1（PARK6）在家族性早发性帕金森病（PD）中经常发生突变，它们在线粒体稳态和线粒体自噬中起着核心作用。当线粒体膜电位去极化时，通过PINK1的作用，帕金被招募到线粒体外膜（MOM），并且能够使线粒体外膜上的孔蛋白、线粒体融合蛋白和米罗蛋白泛素化，然而帕金底物的全部组成——依赖帕金的泛素化修饰组——仍不明确。在此，我们利用定量双甘氨酸捕获蛋白质组学来阐明响应线粒体去极化时依赖帕金的靶标修饰的泛素化位点特异性和拓扑结构。我们鉴定出了数十种蛋白质中数百个动态调节的泛素化位点，其中线粒体外膜蛋白有很强的富集，这表明帕金极大地改变了线粒体蛋白质组的泛素化状态。通过互补的相互作用蛋白质组学，我们发现去极化依赖的帕金与众多线粒体外膜靶标、自噬受体和蛋白酶体相关联。在帕金森病患者中发现发生突变的帕金活性位点残基C431的突变在很大程度上破坏了这些关联。结构和拓扑分析显示，脊椎动物和黑腹果蝇线粒体外膜蛋白的细胞质结构域上依赖帕金的泛素化位点具有广泛的保守性。这些研究为理解PINK1 - 帕金通路如何重塑蛋白质组以支持线粒体稳态提供了资源。
The PARKIN (PARK2) ubiquitin ligase and its regulatory kinase PINK1 (PARK6), often mutated in familial early onset Parkinson’s Disease (PD), play central roles in mitochondrial homeostasis and mitophagy. While PARKIN is recruited to the mitochondrial outer membrane (MOM) upon depolarization via PINK1 action and can ubiquitylate Porin, Mitofusin, and Miro proteins on the MOM, the full repertoire of PARKIN substrates – the PARKIN-dependent ubiquitylome - remains poorly defined. Here we employ quantitative diGLY capture proteomics to elucidate the ubiquitylation site-specificity and topology of PARKIN-dependent target modification in response to mitochondrial depolarization. Hundreds of dynamically regulated ubiquitylation sites in dozens of proteins were identified, with strong enrichment for MOM proteins, indicating that PARKIN dramatically alters the ubiquitylation status of the mitochondrial proteome. Using complementary interaction proteomics, we found depolarization-dependent PARKIN association with numerous MOM targets, autophagy receptors, and the proteasome. Mutation of PARKIN’s active site residue C431, which has been found mutated in PD patients, largely disrupts these associations. Structural and topological analysis revealed extensive conservation of PARKIN-dependent ubiquitylation sites on cytoplasmic domains in vertebrate and D. melanogaster MOM proteins. These studies provide a resource for understanding how the PINK1-PARKIN pathway re-sculpts the proteome to support mitochondrial homeostasis.
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影响因子：
4.5
作者：
Liu S;Sawada T;Lee S;Yu W;Silverio G;Alapatt P;Millan I;Shen A;Saxton W;Kanao T;Takahashi R;Hattori N;Imai Y;Lu B
通讯作者：
Lu B
影响因子：
64.5
作者：
Huttlin EL;Jedrychowski MP;Elias JE;Goswami T;Rad R;Beausoleil SA;Villén J;Haas W;Sowa ME;Gygi SP
通讯作者：
Gygi SP
DOI：
10.1083/jcb.201210111
发表时间：
2013-01-21
期刊：
The Journal of cell biology
影响因子：
--
作者：
Lazarou M;Narendra DP;Jin SM;Tekle E;Banerjee S;Youle RJ
通讯作者：
Youle RJ
影响因子：
13.3
作者：
Narendra, Derek P.;Kane, Lesley A.;Youle, Richard J.
通讯作者：
Youle, Richard J.
影响因子：
8.6
作者：
Dawson, Ted M.;Dawson, Valina L.
通讯作者：
Dawson, Valina L.