The regulation of tau phosphorylation by PCTAIRE 3: implications for the pathogenesis of Alzheimer's disease.
ABSTRACT In the course of Alzheimer's disease, phosphorylated tau aggregates to form paired helical filaments, highly ordered filamentous structures that accumulate within neurons and contribute to the formation of neurofibrillary tangles. This study examines the role of PCTAIRE 3, a cdc2 family protein kinase, within this disease process. We report an elevation in the protein levels of PCTAIRE 3 in the temporal cortex of AD relative to control brains. Analysis of paired helical filaments prepared from AD brain tissue indicates that PCTAIRE 3 is concentrated within this pathological material. Overexpression of PCTAIRE 3 in cell culture suggests that the protein acts indirectly to stimulate phosphorylation at the pT231 and pS235 sites on tau, residues that are modified early in the process of AD pathogenesis. The resurgence of cell cycle proteins is an important mechanism in Alzheimer's disease (AD), and we propose that PCTAIRE 3 is a PHF-associated kinase that modulates tau phosphorylation.
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ABSTRACT: Aggregates of hyperphosphorylated tau protein are found in a group of diseases called tauopathies, which includes Alzheimer's disease. The causes and consequences of tau hyperphosphorylation are routinely investigated in laboratory animals. Mice are the models of choice as they are easily amenable to transgenic technology; consequently, their tau phosphorylation levels are frequently monitored by Western blotting using a panel of monoclonal/polyclonal anti-tau antibodies. Given that mouse secondary antibodies can recognize endogenous mouse immunoglobulins (Igs) and the possible lack of specificity with some polyclonal antibodies, non-specific signals are commonly observed. Here, we characterized the profiles of commonly used anti-tau antibodies in four different mouse models: non-transgenic mice, tau knock-out (TKO) mice, 3xTg-AD mice, and hypothermic mice, the latter a positive control for tau hyperphosphorylation. We identified 3 tau monoclonal antibody categories: type 1, characterized by high non-specificity (AT8, AT180, MC1, MC6, TG-3), type 2, demonstrating low non-specificity (AT270, CP13, CP27, Tau12, TG5), and type 3, with no non-specific signal (DA9, PHF-1, Tau1, Tau46). For polyclonal anti-tau antibodies, some displayed non-specificity (pS262, pS409) while others did not (pS199, pT205, pS396, pS404, pS422, A0024). With monoclonal antibodies, most of the interfering signal was due to endogenous Igs and could be eliminated by different techniques: i) using secondary antibodies designed to bind only non-denatured Igs, ii) preparation of a heat-stable fraction, iii) clearing Igs from the homogenates, and iv) using secondary antibodies that only bind the light chain of Igs. All of these techniques removed the non-specific signal; however, the first and the last methods were easier and more reliable. Overall, our study demonstrates a high risk of artefactual signal when performing Western blotting with routinely used anti-tau antibodies, and proposes several solutions to avoid non-specific results. We strongly recommend the use of negative (i.e., TKO) and positive (i.e., hypothermic) controls in all experiments.PLoS ONE 05/2014; 9(5):e94251. · 3.53 Impact Factor
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ABSTRACT: Background: Alzheimer's disease (AD) is one of the most pressing and difficult to diagnose unmet diseases confronting industrialized countries. It is characterized by the appearance in the post mortem autopsied AD brain of amyloid plaques containing Aβ42 and paired helical filaments in neurofibrillary tangles with hyperphosphorylated tau. Objective: To investigate the potential of proteomics approaches for AD diagnosis. Methods: This reviews focuses on studies of the altered phosphorylation of tau and other proteins as detected in brain biopsy, cerebral spinal fluid (CSF) and blood samples. Results/conclusion: Detection of decreased Aβ42, and increased total and hyperphosphorylated tau in CSF from AD patients can provide a fairly reliable diagnosis. Furthermore, very recent studies have demonstrated altered levels of cytokines in plasma and differential gene expression and protein phosphorylation in peripheral blood mononuclear cells from AD patients. Identification of the roles of these proteins may provide valuable insights into the underlying molecular pathology of AD and possible sites for therapeutic intervention.Expert Opinion on Medical Diagnostics 05/2008; 2(5):577-91.
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ABSTRACT: The objectives of this study were to investigate the protective effects of Eucommia ulmoides Oliv. (EUO) bark and leaf against cytotoxicity induced by amyloid-β peptide (Aβ) and to explore their active components. The PC-12 cells injury mediated by Aβ(25-35) was employed to assess the neuroprotective effects of EUO bark, EUO leaf and various compounds. Intracellular Ca(2+) determination, MTT reduction assay, lactate dehydrogenase leakage evaluation and HO33258/PI staining were used to quantitatively or qualitatively evaluate cell viability and injury. The organic solvents partition and the macroporous resin separation were also applied to tracing the active constituents of EUO bark. Moreover, the effects of 8 compounds (3 iridoid glucoside, 3 phenylpripanoids and 2 flavonoids) were tested to identify the active compounds of EUO leaf. The results demonstrated that the water extracts of EUO barks and leaves, geniposidic acid and chlorogenic acid could efficiently protect PC-12 cells against the cytotoxicity of Aβ(25-35). This research suggests that EUO may represent a potential treatment strategy for Alzheimer's disease. Meanwhile, geniposidic acid and chlorogenic acid are the major active constituents of EUO barks and leaves, respectively.Environmental toxicology and pharmacology. 11/2009; 28(3):342-9.