ALS-linked mutations in ubiquilin-2 or hnRNPA1 reduce interaction between ubiquilin-2 and hnRNPA1
ALS-linked mutations in ubiquilin-2 or hnRNPA1 reduce interaction between ubiquilin-2 and hnRNPA1
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DOI：
10.1093/hmg/ddv020
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发表时间：
2015-05-01
影响因子：
3.5
通讯作者：
Monteiro, Mervyn J.
中科院分区：
文献类型：
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作者：
Gilpin, Kathleen M.;Chang, Lydia;Monteiro, Mervyn J.
肌萎缩侧索硬化症（ALS）相关的突变UBQLN 2和一些成员的异质核核糖核蛋白（hnRNP）家族导致ALS。UBQLN 2中的大多数突变是错义突变，其发生在位于编码蛋白质的中心结构域中的PXX重复基序中和周围。然而，PXX基序的功能和UBQLN 2突变导致ALS的机制都不清楚。我们使用泛素-2的中心结构域筛选酵母双杂交文库，希望鉴定其结合受UBQLN 2突变影响的蛋白。三个这样的相互作用被确定hnRNPA 1，hnRNPA 3和hnRNPU-所有成员的hnRNP家族。这些蛋白质中的每一个中的相互作用区域是它们的富含甘氨酸的结构域，该结构域在引起ALS的hnRNP相关蛋白中最常突变。我们专注于hnRNPA 1，因为蛋白质的突变会导致ALS。我们证实了野生型（WT）泛素-2和hnRNPA 1蛋白在体外和细胞中的相互作用。相比之下，我们检查的泛素-2的5个ALS突变与WT hnRNPA 1的结合减少。此外，携带导致ALS的D262 V错义突变的hnRNPA 1不能结合WT泛素-2。过表达的泛素-2含有ALS突变增加细胞死亡，并为几个突变体，这与增加的hnRNPA 1易位到细胞质。泛素-2的敲低导致hnRNPA 1的周转增加，表明泛素-2的功能是稳定hnRNPA 1。发现泛素-2与hnRNP蛋白相互作用，并且任一蛋白中的突变破坏相互作用，表明蛋白质稳态和RNA代谢之间存在联系。
Amyotrophic lateral sclerosis (ALS)-linked mutations in UBQLN2 and some members of the heterogeneous nuclear ribonucleoproteins (hnRNPs) family cause ALS. Most mutations in UBQLN2 are missense mutations that occur in and around a PXX repeat motif located in the central domain of the encoded protein. However, neither the function of the PXX motif nor the mechanism by which mutations in UBQLN2 cause ALS is known. We screened a yeast two-hybrid library using the central domain of ubiquilin-2 hoping to identify proteins whose binding is affected by the UBQLN2 mutations. Three such interactors were identified-hnRNPA1, hnRNPA3 and hnRNPU-all members of the hnRNP family. The interacting region in each of these proteins was their glycine-rich domain, the domain most frequently mutated in hnRNP-related proteins that cause ALS. We focused on hnRNPA1, because a mutation in the protein causes ALS. We confirmed the interaction between wild-type (WT) ubiquilin-2 and hnRNPA1 proteins in vitro and in cells. In contrast, all five ALS mutations in ubiquilin-2 that we examined had reduced binding with WT hnRNPA1. In addition, hnRNPA1 carrying the D262V missense mutation that causes ALS failed to bind WT ubiquilin-2. Overexpression of ubiquilin-2 containing the ALS mutations increased cell death and, for several of the mutants, this correlated with increased translocation of hnRNPA1 to the cytoplasm. Knockdown of ubiquilin-2 led to increased turnover of hnRNPA1, indicating ubiquilin-2 functions to stabilize hnRNPA1. The discovery that ubiquilin-2 interacts with hnRNP proteins and that mutation in either protein disrupts interaction suggests a connection between proteostasis and RNA metabolism.