Milosch N, Tanriöver G, Kundu A, Rami A, François J-C, Baumkötter F, Weyer S W, Samanta A, Jäschke A, Brod F, Buchholz C J, Kins S, Behl C, Müller U C, Kögel D
Experimental Neurosurgery, Goethe University Hospital, Frankfurt am Main, Germany.
Institute of Cellular and Molecular Anatomy (Anatomie III), Frankfurt University Hospital, Frankfurt am Main, Germany.
Cell Death Dis. 2014 Aug 28;5(8):e1391. doi: 10.1038/cddis.2014.352.
Accumulating evidence indicates that loss of physiologic amyloid precursor protein (APP) function leads to reduced neuronal plasticity, diminished synaptic signaling and enhanced susceptibility of neurons to cellular stress during brain aging. Here we investigated the neuroprotective function of the soluble APP ectodomain sAPPα (soluble APPα), which is generated by cleavage of APP by α-secretase along the non-amyloidogenic pathway. Recombinant sAPPα protected primary hippocampal neurons and SH-SY5Y neuroblastoma cells from cell death induced by trophic factor deprivation. We show that this protective effect is abrogated in neurons from APP-knockout animals and APP-depleted SH-SY5Y cells, but not in APP-like protein 1- and 2- (APLP1 and APLP2) depleted cells, indicating that expression of membrane-bound holo-APP is required for sAPPα-dependent neuroprotection. Trophic factor deprivation diminished the activity of the Akt survival pathway. Strikingly, both recombinant sAPPα and the APP-E1 domain were able to stimulate Akt activity in wild-type (wt) fibroblasts, SH-SY5Y cells and neurons, but failed to rescue in APP-deficient neurons or fibroblasts. The ADAM10 (a disintegrin and metalloproteinase domain-containing protein 10) inhibitor GI254023X exacerbated neuron death in organotypic (hippocampal) slice cultures of wt mice subjected to trophic factor and glucose deprivation. This cell death-enhancing effect of GI254023X could be completely rescued by applying exogenous sAPPα. Interestingly, sAPPα-dependent Akt induction was unaffected in neurons of APP-ΔCT15 mice that lack the C-terminal YENPTY motif of the APP intracellular region. In contrast, sAPPα-dependent rescue of Akt activation was completely abolished in APP mutant cells lacking the G-protein interaction motif located in the APP C-terminus and by blocking G-protein-dependent signaling with pertussis toxin. Collectively, our data provide new mechanistic insights into the physiologic role of APP in antagonizing neurotoxic stress: they suggest that cell surface APP mediates sAPPα-induced neuroprotection via G-protein-coupled activation of the Akt pathway.
越来越多的证据表明，生理性淀粉样前体蛋白（APP）功能丧失会导致神经元可塑性降低、突触信号传导减弱以及大脑衰老过程中神经元对细胞应激的易感性增强。在此，我们研究了可溶性APP胞外结构域sAPPα（可溶性APPα）的神经保护功能，它是由APP通过α-分泌酶沿非淀粉样蛋白生成途径切割产生的。重组sAPPα保护原代海马神经元和SH-SY5Y神经母细胞瘤细胞免受营养因子剥夺诱导的细胞死亡。我们发现，这种保护作用在APP基因敲除动物的神经元和APP缺失的SH-SY5Y细胞中被消除，但在APP样蛋白1和2（APLP1和APLP2）缺失的细胞中未被消除，这表明膜结合型全长APP的表达是sAPPα依赖性神经保护所必需的。营养因子剥夺会降低Akt存活通路的活性。令人惊讶的是，重组sAPPα和APP-E1结构域都能够刺激野生型（wt）成纤维细胞、SH-SY5Y细胞和神经元中的Akt活性，但在APP缺陷的神经元或成纤维细胞中无法发挥挽救作用。ADAM10（一种含解整合素和金属蛋白酶结构域的蛋白10）抑制剂GI254023X加剧了野生型小鼠经营养因子和葡萄糖剥夺处理的器官型（海马）切片培养物中的神经元死亡。通过应用外源性sAPPα可以完全挽救GI254023X的这种细胞死亡增强作用。有趣的是，在缺乏APP细胞内区域C末端YENPTY基序的APP-ΔCT15小鼠的神经元中，sAPPα依赖性Akt诱导不受影响。相反，在缺乏位于APP C末端的G蛋白相互作用基序的APP突变细胞中，以及通过用百日咳毒素阻断G蛋白依赖性信号传导，sAPPα依赖性的Akt激活挽救作用被完全消除。总的来说，我们的数据为APP在对抗神经毒性应激中的生理作用提供了新的机制见解：它们表明细胞表面APP通过G蛋白偶联激活Akt通路介导sAPPα诱导的神经保护作用。
