Capillary Isoelectric Focusing of Akt Isoforms Identifies Highly Dynamic Phosphorylation in Neuronal Cells and Brain Tissue.
Capillary Isoelectric Focusing of Akt Isoforms Identifies Highly Dynamic Phosphorylation in Neuronal Cells and Brain Tissue.
复制标题
DOI：
10.1074/jbc.m115.700138
复制
发表时间：
2016-05-06
期刊：
影响因子：
--
通讯作者：
Eickholt BJ
中科院分区：
文献类型：
--
作者：
Schrötter S;Leondaritis G;Eickholt BJ
PI 3 K/PTEN/Akt通路已被确立为核心信号通路，其对于神经元整合到神经元回路中以及维持成人脑中神经元的结构和功能至关重要。Akt 1 -3激酶在生长因子信号传导后通过残基Thr 308和Ser 473上的两个磷酸化事件特异性激活，其随后磷酸化大量下游靶点。然而，我们仍然缺乏对PI 3 K/PTEN/Akt通路中亚型特异性的复杂性和调节的清楚理解。我们利用基于毛细管的等电聚焦方法来研究神经元细胞和发育中的大脑中Akt磷酸化的动力学，并确定以前未描述的Akt磷酸化和激活的特征。首先，我们表明，在急性PI 3 K激活时，Akt形式上多种磷酸化事件的积累与Ser 473和Thr 308磷酸化同时发生，并为Ser 473和Thr 308磷酸化的解偶联以及Akt 1形式对PI 3 K抑制的敏感性差异提供了证据。其次，我们检测到一个短暂的转变Akt亚型磷酸化和激活模式在出生后早期的大脑发育，在相应的突触发育和成熟的阶段。第三，我们显示了Ser 473-Akt种类对成熟神经元中的PTEN缺失的不同敏感性，这表明与由PTEN控制的库相比，可接近生长因子的Akt库中存在固有差异。我们的研究表明，在神经元中存在时间和信号依赖性的Akt的复杂磷酸化事件。
The PI3K/PTEN/Akt pathway has been established as a core signaling pathway that is crucial for the integration of neurons into neuronal circuits and the maintenance of the architecture and function of neurons in the adult brain. Akt1–3 kinases are specifically activated by two phosphorylation events on residues Thr308 and Ser473 upon growth factor signaling, which subsequently phosphorylate a vast cohort of downstream targets. However, we still lack a clear understanding of the complexity and regulation of isoform specificity within the PI3K/PTEN/Akt pathway. We utilized a capillary-based isoelectric focusing method to study dynamics of Akt phosphorylation in neuronal cells and the developing brain and identify previously undescribed features of Akt phosphorylation and activation. First, we show that the accumulation of multiple phosphorylation events on Akt forms occur concurrently with Ser473 and Thr308 phosphorylation upon acute PI3K activation and provide evidence for uncoupling of Ser473 and Thr308 phosphorylation, as well as differential sensitivities of Akt1 forms upon PI3K inhibition. Second, we detect a transient shift in Akt isoform phosphorylation and activation pattern during early postnatal brain development, at stages corresponding to synapse development and maturation. Third, we show differential sensitivities of Ser473-Akt species to PTEN deletion in mature neurons, which suggests inherent differences in the Akt pools that are accessible to growth factors as compared with the pools that are controlled by PTEN. Our study demonstrates the presence of complex phosphorylation events of Akt in a time- and signal-dependent manner in neurons.