14-3-3 protein and gene profiling in epileptic brain

描述(由申请人提供)：癫痫发作可导致患者进行性神经变性和认知障碍，与癫痫持续状态无关。使用一种新的小鼠局灶性惊厥模型和脑电定量，我们已经从耐受性发作中分离出损伤性惊厥的脑电(EEG)特征。其次，我们已经确定了14-3-3分子伴侣蛋白家族在癫痫发作后细胞死亡途径的协调中的重要作用，并证明了与14-3-3相互作用是识别癫痫发作后损伤的新的介质的有效预测工具。 需要检验的假说是：产生损伤的癫痫和提供损伤保护的癫痫引起独特的基因组指纹，这些指纹编码与细胞死亡调节因子14-3-3家族相互作用的新蛋白，从而产生新的靶向调控因子，调节癫痫发作后细胞死亡和存活的平衡。具体目标是： 1)通过比较非损伤性惊厥动物海马神经元的基因组反应和非损伤性惊厥后的反应，以及对损伤性惊厥的耐受性，我们将找出预测惊厥后结局的特定遗传标记； 2)利用14-3-3蛋白作为重点功能筛选，在调控基因中寻找与控制癫痫诱导的细胞死亡最相关的基因。 我们的实验室致力于了解癫痫患者癫痫发作、脑损伤和细胞死亡途径之间的关系，这些细胞死亡途径是癫痫患者神经变性的基础。我们集中应用DNA微阵列来分离特定癫痫发作的基因组指纹，为我们提供了一个追求这一目标的新工具；将其与基于功能的筛选工具(与14-3-3蛋白质的相互作用)相结合，将建立一种新的方法，将微阵列数据置于可用的背景下。这一R21建议具有重要的实际应用；这些数据将在可识别的癫痫类型的背景下建立一个细胞死亡调节基因队列，使我们能够对导致脑损伤的癫痫的后果和治疗有新的见解。

DESCRIPTION (provided by applicant): Epileptic seizures can cause progressive neurodegeneration and cognitive impairment in patients independent of episodes of status epilepticus. Using a novel mouse model of focally-evoked seizures with EEG quantification we have isolated the electrographic (EEG) signature of injury-causing seizures from seizures that are tolerance-inducing. Second, we have identified a preeminent role for the 14-3-3 family of molecular chaperone proteins in the coordination of cell death pathways after seizures and we demonstrate that interaction with 14-3-3 is a potent predictive tool for identifying novel mediators of injury following seizures. The hypothesis to be tested is: injury-producing seizures and seizures that confer protection against injury evoke unique genomic fingerprints which encode novel proteins that interact with the 14-3-3 family of cell death regulators, thus yielding new target regulators of the balance of cell death and survival after seizures. The specific aims are: 1) By contrasting the genomic response of hippocampal neurons from animals subject to injury-producing seizures with the response following non-injury producing seizures and seizures that confer tolerance against injury-producing seizures we will identify specific genetic markers that predict outcome following seizures; 2) To use 14-3-3 proteins as a focused functional screen to identify among the regulated genes those most relevant to the control of seizure-induced cell death. Our laboratory is focused on understanding the relationship between seizures, brain damage and the cell death pathways that underlie neurodegeneration in epilepsy patients. Our focused application of DNA microarrays to isolate the genomic fingerprint of specific epileptic seizures provides us with a new tool in the pursuit of this goal; combining this with a function-based screening tool (interaction with 14-3-3 proteins) will establish a new approach placing microarray data into a useable context. This R21 proposal has significant practical applications; the data will establish a cohort of cell death-regulating genes in the context of identifiable seizure types, endowing us with new insight into the consequences and treatment of those seizures resulting in brain injury.