Novel α-secretase cleavage of Alzheimer's amyloid β precursor protein in the endoplasmic reticulum of COS7 cells
Novel α-secretase cleavage of Alzheimer's amyloid β precursor protein in the endoplasmic reticulum of COS7 cells
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DOI：
10.1016/j.neulet.2004.11.032
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发表时间：
2005-03
影响因子：
2.5
通讯作者：
R. Shin;T. Saido;M. Maeda;T. Kitamoto
中科院分区：
文献类型：
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作者：
R. Shin;T. Saido;M. Maeda;T. Kitamoto
在APP的加工过程中，α-和β-分泌酶途径相互竞争以切割APP。因此，生理上这两种分泌酶可能在相同的亚细胞区室中共定位。以前发现β分泌酶在内质网（ER）中切割APP。我们在此测试α-分泌酶的切割是否也在内质网中检测到。我们采用转染人APP695 cDNA的COS7细胞实验体系，对细胞裂解液和培养基进行蛋白水解产物检测。当通过bfa介导的转运抑制或使用含有ER检索基序的突变APP将APP表达集中在内质网中时，α-分泌酶产物sAPPα在细胞中积累。免疫荧光显微镜显示，内质网靶向的APP与内质网标记物共定位，产生细胞内sAPPα的积累。这些结果表明α-分泌酶在内质网中发生了APP的裂解。我们进一步研究了PKC的直接激活剂phbol酯PDBu对内质网中APP α-分泌酶和β-分泌酶裂解的影响。PDBu处理转染er靶向APP的COS7细胞增加了sAPPα的产生，反过来减少了β分泌酶产物sAPPβ的产生。因此，在内质网中，α-分泌酶与β-分泌酶竞争裂解APP，这种竞争相关性可能调节了在内质网中发现的a - β42的产生。
In the processing of APP, α- and β-secretase pathways compete with each other for cleaving APP. Therefore, physiologically these two secretases are likely to colocalize in the same subcellular compartments. Previously β-secretase cleavage of APP was found in the endoplasmic reticulum (ER). We herein tested whether α-secretase cleavage is also detected in the ER. We used experimental system of COS7 cells transfected with cDNA encoding human APP695, and the cell lysates and media were examined for its proteolytic products. When APP expression is concentrated in the ER by BFA-mediated transport inhibition or by using mutant APP harboring an ER-retrieval motif, α-secretase product sAPPα was accumulated in the cells. Immunofluorescence microscopy revealed that the ER-targeted APP produced intracellular accumulation of sAPPα, colocalizing with an ER marker. These results indicate that α-secretase cleavage of APP occurs in the ER. Further we examined the effects of phorbol ester PDBu, a direct activator of PKC, on the α-secretase and β-secretase cleavages of APP occurring in the ER. Treatment with PDBu of COS7 cells transfected with the ER-targeted APP increased production of sAPPα and conversely decreased production of β-secretase product sAPPβ. Thus, in the ER, α-secretase competes with β-secretase for cleaving APP and such competitive correlation might modulate the production of Aβ42 found in this compartment.