Merrick S E, Demoise D C, Lee V M
David Mahoney Institute of Neurological Sciences and the Department of Pathology and Laboratory Medicine, University of Pennsylvania School of Medicine, Philadelphia, 19104-4283, USA.
J Biol Chem. 1996 Mar 8;271(10):5589-94. doi: 10.1074/jbc.271.10.5589.
Tau proteins isolated from paired helical filaments, the major building blocks of Alzheimer's disease neurofibrillary tangle, are abnormally phosphorylated and unable to bind microtubules. To examine the dynamics of tau phosphorylation and to identify specific tau phosphorylation sites involved in the stabilization of microtubules, we treated cultured postmitotic neuron-like cells (NT2N) derived from a human teratocarcinoma cell line (NTera2/D1) with drugs that depolymerize microtubules (i.e. colchicine or nocodazole). This led to the recovery of dephosphorylated tau from the NT2N cells as monitored by a relative increase in the electrophoretic mobility of tau and an increase in the turnover of [32P]PO4-labeled tau. However, not all phosphorylation sites on tau are affected by colchicine or nocodazole. Ser202/Thr205 appears to be completely and specifically dephosphorylated by protein phosphatase 2A since this dephosphorylation was blocked by inhibitors of protein phosphatase 2A but not by inhibitors of protein phosphatase 2B. These findings, together with the recent observation that protein phosphatase 2A is normally bound to microtubules in intact cells, suggest that the polymerization state of microtubules could modulate the phosphorylation state of tau at specific sites in the normal and Alzheimer's disease brain.
从成对螺旋丝中分离出的tau蛋白是阿尔茨海默病神经原纤维缠结的主要组成部分，其磷酸化异常且无法结合微管。为了研究tau蛋白磷酸化的动力学，并确定参与微管稳定的特定tau蛋白磷酸化位点，我们用能使微管解聚的药物（即秋水仙碱或诺考达唑）处理源自人畸胎瘤细胞系（NTera2/D1）的有丝分裂后神经元样细胞（NT2N）。通过tau蛋白电泳迁移率的相对增加以及[32P]PO4标记的tau蛋白周转率的增加监测发现，这导致NT2N细胞中去磷酸化的tau蛋白恢复。然而，tau蛋白上并非所有磷酸化位点都受秋水仙碱或诺考达唑影响。Ser202/Thr205似乎被蛋白磷酸酶2A完全且特异性地去磷酸化，因为这种去磷酸化被蛋白磷酸酶2A抑制剂阻断，但未被蛋白磷酸酶2B抑制剂阻断。这些发现，连同最近观察到蛋白磷酸酶2A在完整细胞中通常与微管结合，表明微管的聚合状态可能在正常和阿尔茨海默病大脑中调节特定位点tau蛋白的磷酸化状态。
