Interaction between AP-5 and the hereditary spastic paraplegia proteins SPG11 and SPG15.
Interaction between AP-5 and the hereditary spastic paraplegia proteins SPG11 and SPG15.
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DOI：
10.1091/mbc.e13-03-0170
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发表时间：
2013-08
影响因子：
3.3
通讯作者：
Robinson MS
中科院分区：
文献类型：
--
作者：
Hirst J;Borner GH;Edgar J;Hein MY;Mann M;Buchholz F;Antrobus R;Robinson MS
接头蛋白复合物AP-5与遗传性痉挛性截瘫蛋白SPG 11和SPG 15以明显等摩尔的比例稳定地结合，形成一种新型的外壳。结果表明，SPG 15有利于对接的外套到晚期内体/溶酶体膜和SPG 11（可能与SPG 15）作为一个支架。AP-5复合物是异源四聚体衔接蛋白（AP复合物）家族中最近发现的但在进化上古老的成员。它与遗传性痉挛性截瘫患者中突变的两种蛋白质SPG 11和SPG 15相关。在这里，我们表明，AP-5的四个亚基可以与SPG 11和SPG 15，无论是从细胞质和洗涤剂提取的膜免疫共沉淀，化学计量比为100：1：1：1：1：1。AP-5亚基的SPG 11或SPG 15表型敲除：所有六种敲除均导致阳离子非依赖性甘露糖6-磷酸受体被捕获在早期内体簇中。此外，AP-5、SPG 11和SPG 15共定位于晚期内体/溶酶体区室。SPG 11和SPG 15都预测了含有与网格蛋白重链和COPI亚基相关的α-β的二级结构。SPG 11还具有N-末端β-螺旋桨样结构域，其在体外与AP-5相互作用。我们认为AP-5、SPG 15和SPG 11形成了一个外套样复合物，其中AP-5参与蛋白分选，SPG 15通过其FYVE结构域与PI 3 P相互作用促进外套在膜上的对接，SPG 11（可能与SPG 15一起）形成了一个支架。
The adaptor protein complex AP-5 is stably associated with the hereditary spastic paraplegia proteins SPG11 and SPG15 in an apparently equimolar ratio to form a new type of coat. Results suggest that SPG15 facilitates the docking of the coat onto late endosomal/lysosomal membranes and that SPG11 (possibly together with SPG15) acts as a scaffold. The AP-5 complex is a recently identified but evolutionarily ancient member of the family of heterotetrameric adaptor proteins (AP complexes). It is associated with two proteins that are mutated in patients with hereditary spastic paraplegia, SPG11 and SPG15. Here we show that the four AP-5 subunits can be coimmunoprecipitated with SPG11 and SPG15, both from cytosol and from detergent-extracted membranes, with a stoichiometry of ∼1:1:1:1:1:1. Knockdowns of SPG11 or SPG15 phenocopy knockdowns of AP-5 subunits: all six knockdowns cause the cation-independent mannose 6-phosphate receptor to become trapped in clusters of early endosomes. In addition, AP-5, SPG11, and SPG15 colocalize on a late endosomal/lysosomal compartment. Both SPG11 and SPG15 have predicted secondary structures containing α-solenoids related to those of clathrin heavy chain and COPI subunits. SPG11 also has an N-terminal, β-propeller–like domain, which interacts in vitro with AP-5. We propose that AP-5, SPG15, and SPG11 form a coat-like complex, with AP-5 involved in protein sorting, SPG15 facilitating the docking of the coat onto membranes by interacting with PI3P via its FYVE domain, and SPG11 (possibly together with SPG15) forming a scaffold.