Efthimiopoulos S, Vassilacopoulou D, Ripellino J A, Tezapsidis N, Robakis N K
Department of Psychiatry and Fishberg Research Center for Neurobiology, Mount Sinai School of Medicine, NY 10029, USA.
Proc Natl Acad Sci U S A. 1996 Jul 23;93(15):8046-50. doi: 10.1073/pnas.93.15.8046.
The Abeta peptide of Alzheimer disease is derived from the proteolytic processing of the amyloid precursor proteins (APP), which are considered type I transmembrane glycoproteins. Recently, however, soluble forms of full-length APP were also detected in several systems including chromaffin granules. In this report we used antisera specific for the cytoplasmic sequence of APP to show that primary bovine chromaffin cells secrete a soluble APP, termed solAPPcyt, of an apparent molecular mass of 130 kDa. This APP was oversecreted from Chinese hamster ovary cells transfected with a full-length APP cDNA indicating that solAPPcyt contained both the transmembrane and Abeta sequence. Deglycosylation of solAPPcyt showed that it contained both N- and O-linked sugars, suggesting that this APP was transported through the endoplasmic reticulum-Golgi pathway. Secretion of solAPPcyt from primary chromatin cells was temperature-, time-, and energy-dependent and was stimulated by cell depolarization in a Ca2+-dependent manner. Cholinergic receptor agonists, including acetylcholine, nicotine, or carbachol, stimulated the rapid secretion of solAPPcyt, a process that was inhibited by cholinergic antagonists. Stimulation of solAPPcyt secretion was paralleled by a stimulation of secretion in catecholamines and chromogranin A, indicating that secretion of solAPPcyt was mediated by chromaffin granule vesicles. Taken together, our results show that release of the potentially amyloidogenic solAPPcyt is an active cellular process mediated by both the constitutive and regulated pathways. solAPPcyt was also detected in human cerebrospinal fluid. Combined with the neuronal physiology of chromaffin cells, our data suggest that cholinergic agonists may stimulate the release of this APP in neuronal synapses where it may exert its biological functions. Moreover, vesicular or secreted solAPPcyt may serve as a soluble precursor of Abeta.
阿尔茨海默病的β-淀粉样肽源自淀粉样前体蛋白（APP）的蛋白水解过程，淀粉样前体蛋白被认为是I型跨膜糖蛋白。然而，最近在包括嗜铬颗粒在内的多个系统中也检测到了全长APP的可溶性形式。在本报告中，我们使用针对APP细胞质序列的抗血清来表明，原代牛嗜铬细胞分泌一种表观分子量为130 kDa的可溶性APP，称为solAPPcyt。这种APP从转染了全长APP cDNA的中国仓鼠卵巢细胞中过度分泌，表明solAPPcyt包含跨膜和β-淀粉样序列。solAPPcyt的去糖基化表明它同时含有N-连接和O-连接糖，这表明这种APP是通过内质网-高尔基体途径运输的。solAPPcyt从原代嗜铬细胞中的分泌是温度、时间和能量依赖性的，并以Ca2+依赖性方式受到细胞去极化的刺激。胆碱能受体激动剂，包括乙酰胆碱、尼古丁或卡巴胆碱，刺激solAPPcyt的快速分泌，这一过程被胆碱能拮抗剂抑制。solAPPcyt分泌的刺激与儿茶酚胺和嗜铬粒蛋白A分泌的刺激平行，表明solAPPcyt的分泌是由嗜铬颗粒小泡介导的。综上所述，我们的结果表明，潜在的淀粉样生成性solAPPcyt的释放是一个由组成型和调节型途径介导的活跃细胞过程。在人类脑脊液中也检测到了solAPPcyt。结合嗜铬细胞的神经元生理学，我们的数据表明胆碱能激动剂可能在神经元突触中刺激这种APP的释放，在那里它可能发挥其生物学功能。此外，囊泡性或分泌性solAPPcyt可能作为β-淀粉样肽的可溶性前体。
