Paul Ehrlich Institute, Paul Ehrlich-Straße 51-59, D-63225 Langen, Germany.
Cell Commun Signal. 2010 Sep 28;8:28. doi: 10.1186/1478-811X-8-28.
Prion diseases or transmissible spongiform encephalopathies (TSEs) are fatal diseases associated with the conversion of the cellular prion protein (PrPC) to the abnormal prion protein (PrPSc). Since the molecular mechanisms in pathogenesis are widely unclear, we analyzed the global phospho-proteome and detected a differential pattern of tyrosine- and threonine phosphorylated proteins in PrPSc-replicating and pentosan polysulfate (PPS)-rescued N2a cells in two-dimensional gel electrophoresis. To quantify phosphorylated proteins, we performed a SILAC (stable isotope labeling by amino acids in cell culture) analysis and identified 105 proteins, which showed a regulated phosphorylation upon PrPSc infection. Among those proteins, we validated the dephosphorylation of stathmin and Cdc2 and the induced phosphorylation of cofilin in PrPSc-infected N2a cells in Western blot analyses. Our analysis showed for the first time a differentially regulated phospho-proteome in PrPSc infection, which could contribute to the establishment of novel protein markers and to the development of novel therapeutic intervention strategies in targeting prion-associated disease.
朊病毒病或传染性海绵状脑病（TSE）是与细胞朊蛋白（PrPC）转化为异常朊蛋白（PrPSc）相关的致命疾病。由于发病机制中的分子机制尚不清楚，我们分析了全球磷酸化蛋白质组，并在二维凝胶电泳中检测到 PrPSc 复制和戊聚糖多硫酸酯（PPS）挽救的 N2a 细胞中酪氨酸和苏氨酸磷酸化蛋白质的差异模式。为了定量磷酸化蛋白质，我们进行了 SILAC（细胞培养中的氨基酸稳定同位素标记）分析，鉴定了 105 种蛋白质，这些蛋白质在 PrPSc 感染时表现出调节性磷酸化。在这些蛋白质中，我们通过 Western blot 分析验证了 PrPSc 感染的 N2a 细胞中 stathmin 和 Cdc2 的去磷酸化以及 cofilin 的诱导磷酸化。我们的分析首次显示了 PrPSc 感染中差异调节的磷酸化蛋白质组，这可能有助于建立新的蛋白质标志物，并开发针对朊病毒相关疾病的新的治疗干预策略。
