Riefler Gary M, Balasingam Gaithri, Lucas Kenyatta G, Wang Sheng, Hsu Shu-Chan, Firestein Bonnie L
Department of Cell Biology and Neuroscience, Rutgers, The State University of New Jersey, Nelson Biological Laboratories, 604 Allison Road, Piscataway, NJ 08854-8082, USA.
Biochem J. 2003 Jul 1;373(Pt 1):49-55. doi: 10.1042/BJ20021838.
The PDZ domains of postsynaptic density (PSD) protein-95 play a role in the localization of PSD-95 and binding partners to neuronal synapses. The identification of binding partners to these PDZ domains can help us in understanding how signalling complexes are assembled. We observed that one of the subunits in the sec6/8 or exocyst complex, sec8, contains a C-terminal consensus sequence for PDZ binding. Sec8 binds to PDZ1-2 of PSD-95, and this binding can be competed with a peptide that binds to PDZ1 and PDZ2 in the peptide-binding site. In addition, binding of sec8 is dependent on its C-terminal-binding sequence namely Thr-Thr-Val (TTV). Immunoblotting of rat tissue extracts shows that sec8 and PSD-95 are enriched in the same brain regions, and sec8 and PSD-95 have the same subcellular distribution in pheochromocytoma cells, suggesting that these proteins may interact in vivo. Immunoprecipitation studies of sec8 and PSD-95 in brain provide further evidence of a sec8 and PSD-95 interaction. Furthermore, the cytosolic PSD-95 interactor competes with sec8 for interaction with PSD-95. Taken together, our results suggest that the cytosolic PSD-95 interactor may function to regulate the ability of sec8 to bind to PSD-95.
突触后致密蛋白（PSD）-95的PDZ结构域在PSD-95及其结合伴侣定位于神经元突触的过程中发挥作用。鉴定这些PDZ结构域的结合伴侣有助于我们理解信号复合物是如何组装的。我们观察到，sec6/8或外泌体复合物中的一个亚基sec8含有一个用于PDZ结合的C末端共有序列。Sec8与PSD-95的PDZ1-2结合，这种结合可以被一种在肽结合位点与PDZ1和PDZ2结合的肽所竞争。此外，sec8的结合依赖于其C末端结合序列，即苏氨酸-苏氨酸-缬氨酸（TTV）。对大鼠组织提取物的免疫印迹显示，sec8和PSD-95在相同的脑区富集，并且sec8和PSD-95在嗜铬细胞瘤细胞中具有相同的亚细胞分布，这表明这些蛋白质可能在体内相互作用。对大脑中sec8和PSD-95的免疫沉淀研究为sec8和PSD-95的相互作用提供了进一步的证据。此外，胞质PSD-95相互作用蛋白与sec8竞争与PSD-95的相互作用。综上所述，我们的结果表明，胞质PSD-95相互作用蛋白可能起到调节sec8与PSD-95结合能力的作用。
