Adaptor protein complex 4 (AP-4) is a heterotetrameric complex that promotes export of selected cargo proteins from the -Golgi network. Mutations in each of the AP-4 subunits cause a complicated form of Hereditary Spastic Paraplegia (HSP). Herein, we report that ApoER2, a receptor in the Reelin signaling pathway, is a cargo of the AP-4 complex. We identify the motif ISSF/Y within the ApoER2 cytosolic domain as necessary for interaction with the canonical signal-binding pocket of the µ4 (AP4M1) subunit of AP-4. - knock-out (KO) HeLa cells and hippocampal neurons from -KO mice display increased co-localization of ApoER2 with Golgi markers. Furthermore, hippocampal neurons from -KO mice and -KO human iPSC-derived cortical i3Neurons exhibit reduced ApoER2 protein expression. Analyses of biosynthetic transport of ApoER2 reveal differential post-Golgi trafficking of the receptor, with lower axonal distribution in KO compared to wild-type neurons, indicating a role of AP-4 and the ISSF/Y motif in the axonal localization of ApoER2. Finally, analyses of Reelin signaling in mouse hippocampal and human cortical KO neurons show that AP4 deficiency causes no changes in Reelin-dependent activation of the AKT pathway and only mild changes in Reelin-induced dendritic arborization, but reduces Reelin-induced ERK phosphorylation, CREB activation, and Golgi deployment. This work thus establishes ApoER2 as a novel cargo of the AP-4 complex, suggesting that defects in the trafficking of this receptor and in the Reelin signaling pathway could contribute to the pathogenesis of HSP caused by mutations in AP-4 subunits.

中文翻译：

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Reelin 受体 ApoER2 是衔接蛋白复合物 AP-4 的货物：对遗传性痉挛性截瘫的影响

衔接蛋白复合物 4 (AP-4) 是一种异四聚体复合物，可促进高尔基体网络中选定的货物蛋白的输出。每个 AP-4 亚基的突变都会导致一种复杂的遗传性痉挛性截瘫 (HSP)。在此，我们报道 Reelin 信号通路中的受体 ApoER2 是 AP-4 复合物的货物。我们鉴定出 ApoER2 胞质结构域内的基序 ISSF/Y 是与 AP-4 µ4 (AP4M1) 亚基的典型信号结合袋相互作用所必需的。 - 敲除 (KO) 小鼠的 HeLa 细胞和海马神经元显示 ApoER2 与高尔基体标记的共定位增加。此外，来自 -KO 小鼠和 -KO 人 iPSC 衍生的皮质 i3Neurons 的海马神经元表现出 ApoER2 蛋白表达减少。对 ApoER2 生物合成转运的分析揭示了受体的高尔基体后转运差异，与野生型神经元相比，KO 中的轴突分布较低，表明 AP-4 和 ISSF/Y 基序在 ApoER2 轴突定位中的作用。最后，对小鼠海马和人类皮质 KO 神经元中 Reelin 信号传导的分析表明，AP4 缺陷不会导致 Reelin 依赖性 AKT 通路激活发生变化，仅导致 Reelin 诱导的树突状树枝化发生轻微变化，但会降低 Reelin 诱导的 ERK 磷酸化、CREB激活和高尔基体部署。因此，这项工作将 ApoER2 确立为 AP-4 复合物的新货物，表明该受体运输和 Reelin 信号通路中的缺陷可能导致 AP-4 亚基突变引起的 HSP 发病机制。