Retromer binding to FAM21 and the WASH complex is perturbed by the Parkinson disease-linked VPS35(D620N) mutation.
Retromer binding to FAM21 and the WASH complex is perturbed by the Parkinson disease-linked VPS35(D620N) mutation.
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DOI：
10.1016/j.cub.2014.06.024
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发表时间：
2014-07-21
期刊：
影响因子：
9.2
通讯作者：
Cullen, Peter J.
中科院分区：
文献类型：
--
作者：
McGough, Ian J.;Steinberg, Florian;Jia, Da;Barbuti, Peter A.;McMillan, Kirsty J.;Heesom, Kate J.;Whone, Alan L.;Caldwell, Maeve A.;Billadeau, Daniel D.;Rosen, Michael K.;Cullen, Peter J.
逆转录聚体是一种蛋白质组装体，它在协调跨膜货物蛋白从内体输出到定位于高尔基体和质膜的检索途径中发挥核心作用。最近，逆转录体成分VPS35的一个特殊突变VPS35(D620N)将逆转录反应障碍与家族性常染色体显性遗传和散发性帕金森病联系在一起。然而，这种突变对逆转录功能的影响仍然没有得到很好的描述。在这里，我们证实了在表达VPS35(D620N)的细胞中，内小体到TGN的转运存在扰动，但内小体到质膜的循环没有扰动，我们在携带VPS35(D620N)突变的患者细胞中证实了这一点。通过基于细胞培养氨基酸比较稳定同位素标记(SILAC)的野生型VPS35与VPS35(D620N)突变相互作用的分析，我们确定D620N突变的主要缺陷在于与肌动蛋白核Wiskott-Aldrich综合征和SCAR同源(WASH)复合体的关联。此外，利用等温量热法，我们确定VPS35(D620N)突变体的主要缺陷是与洗涤复合体成分FAM21的亲和力降低2.2±0.5倍。这些数据定义了由VPS35(D620N)突变引起的逆转录病毒组装中的主要分子缺陷，并通过揭示逆转录病毒介导的内小体到TGN运输的功能效应，为帕金森病的逆转录病毒调控解除提供了新的见解。VPS35(D620N)突变导致内小体到TGN的运输缺陷突变与肌动蛋白聚合WASH复合体的关联受到干扰原始缺陷是结合WASH组件的亲和力降低McGough等人利用定量蛋白质组学和患者来源的成纤维细胞的功能分析为帕金森病逆转录聚合放松调控提供了新的分子见解。确定与帕金森病相关的逆转录病毒VPS35(D620N)突变相关的主要分子缺陷，并通过揭示逆转录病毒介导的内小体到TGN转运的影响，为研究逆转录病毒对帕金森病的解除调控提供新的见解。
Retromer is a protein assembly that plays a central role in orchestrating export of transmembrane-spanning cargo proteins from endosomes into retrieval pathways destined for the Golgi apparatus and the plasma membrane. Recently, a specific mutation in the retromer component VPS35, VPS35(D620N), has linked retromer dysfunction to familial autosomal dominant and sporadic Parkinson disease. However, the effect of this mutation on retromer function remains poorly characterized. Here we established that in cells expressing VPS35(D620N) there is a perturbation in endosome-to-TGN transport but not endosome-to-plasma membrane recycling, which we confirm in patient cells harboring the VPS35(D620N) mutation. Through comparative stable isotope labeling by amino acids in cell culture (SILAC)-based analysis of wild-type VPS35 versus the VPS35(D620N) mutant interactomes, we establish that the major defect of the D620N mutation lies in the association to the actin-nucleating Wiskott-Aldrich syndrome and SCAR homolog (WASH) complex. Moreover, using isothermal calorimetry, we establish that the primary defect of the VPS35(D620N) mutant is a 2.2 ± 0.5-fold decrease in affinity for the WASH complex component FAM21. These data define the primary molecular defect in retromer assembly that arises from the VPS35(D620N) mutation and, by revealing functional effects on retromer-mediated endosome-to-TGN transport, provide new insight into retromer deregulation in Parkinson disease. VPS35(D620N) mutation leads to a defect in endosome-to-TGN transport Mutant has a perturbed association with the actin-polymerizing WASH complex Primary defect is a reduced affinity for binding WASH component FAM21 Provides new molecular insight into retromer deregulation in Parkinson disease Using quantitative proteomics and functional analysis in patient-derived fibroblasts, McGough et al. define the primary molecular defect associated with the Parkinson disease-linked retromer VPS35(D620N) mutation and, by revealing effects on retromer-mediated endosome-to-TGN transport, provide new insight into retromer deregulation in this disease.
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