Demonstration of BACE (beta-secretase) phosphorylation and its interaction with GGA1 in cells by fluorescence-lifetime imaging microscopy.
Demonstration of BACE (beta-secretase) phosphorylation and its interaction with GGA1 in cells by fluorescence-lifetime imaging microscopy.
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发表时间：
2004
影响因子：
4
通讯作者：
C. V. von Arnim;Michelle M. Tangredi;I. Peltan;Bonny M. Lee;M. Irizarry;A. Kinoshita;B. Hyman
中科院分区：
文献类型：
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作者：
C. V. von Arnim;Michelle M. Tangredi;I. Peltan;Bonny M. Lee;M. Irizarry;A. Kinoshita;B. Hyman
β -分泌酶（BACE）执行两个蛋白水解步骤中的第一个步骤，生成淀粉样β肽，这些淀粉样β肽积聚在阿尔茨海默病（AD）的老年斑中。由于大多数BACE活动发生在核内体中，因此调节其运输到这些腔室的信号对于了解AD的发病机制非常重要。BACE c端附近的DISLL序列介导BACE与高尔基定位的含γ -耳arf结合（GGA）蛋白的VHS结构域的结合，参与蛋白质向内体的分选。基序丝氨酸残基的磷酸化调节BACE从早期内体再循环回细胞表面，并在分离蛋白检测中增强BACE与GGA蛋白的相互作用。我们发现BACE磷酸化影响细胞中BACE- gga相互作用，使用一种新的基于荧光共振能量转移的蛋白质接近性，荧光寿命成像。虽然丝氨酸磷酸化的BACE分布在整个细胞中，但GGA1与野生型蛋白的相互作用发生在核旁室。假磷酸化和非磷酸化的BACE突变体在高尔基体中仍然定位于GGA1，但后者的突变减弱了这两种蛋白的FRET信号。由于在人脑中可以鉴定出丝氨酸残基磷酸化的BACE，这些数据表明BACE的丝氨酸磷酸化是一种生理上相关的翻译后修饰，通过与GGA1相互作用调节核旁室的运输。
beta-Secretase (BACE) carries out the first of two proteolysis steps to generate the amyloid-beta peptides that accumulate in the senile plaques in Alzheimer's disease (AD). Because most BACE activity occurs in endosomes, signals regulating its trafficking to these compartments are important to an understanding of AD pathogenesis. A DISLL sequence near the BACE C-terminus mediates binding of BACE to the VHS domains of Golgi-localized gamma-ear-containing ARF-binding (GGA) proteins, which are involved in the sorting of proteins to endosomes. Phosphorylation of the motif's serine residue regulates BACE recycling back to the cell surface from early endosomes and enhances the interaction of BACE with GGA proteins in isolated protein assays. We found that BACE phosphorylation influences BACE-GGA interactions in cells using a new fluorescence-resonance-energy-transfer-based assay of protein proximity, fluorescence lifetime imaging. Although serine-phosphorylated BACE was distributed throughout the cell, interaction of GGA1 with the wild-type protein occurred in juxtanuclear compartments. Pseudo-phosphorylated and non-phosphorylated BACE mutants remained localized with GGA1 in the Golgi body, but the latter mutation diminished the two proteins' FRET signal. Because BACE phosphorylated at serine residues can be identified in human brain, these data suggest that serine phosphorylation of BACE is a physiologically relevant post-translational modification that regulates trafficking in the juxtanuclear compartment by interaction with GGA1.