PROLINE DIRECTED KINASES IN ALZHEIMERS DISEASE

这项建议的长期目标是了解病因， 阿尔茨海默病（AD）的发病机制。 越来越多的证据表明， 在AD中，几种神经元蛋白的磷酸化状态改变。 这种异常的磷酸化作用显然会导致不同的 细胞功能，从而最终导致神经元死亡。 特别是 已证明微管相关蛋白tau存在于 AD的成对螺旋丝（PHF）中的过度磷酸化状态 个脑袋 最近的证据表明PHF tau蛋白的过度磷酸化 可以在很大程度上被催化。但不限于脯氨酸导向蛋白 激酶（PDPK）。有三种众所周知的PDPK：MAP激酶，cdc 2激酶， 和糖原合成酶激酶3。 PDPK和非PDPK的身份 目前，负责PHF tau在体内过度磷酸化的是 不清楚本建议的具体目的是：1）比较 tau蛋白被三种PDPK磷酸化。 磷酸化动力学 并且通过肽图谱比较tau上磷酸化的位点， 通过不同的磷酸化位点特异性抗tau抗体; 2） 研究非PDPK在tau过度磷酸化中的作用。 六非 PDPK将用于制备六种磷酸化tau。 的 将比较这些tau种类被PDPK进一步磷酸化的情况; 3)使用不同的合成肽含有已知的磷酸化 tau的位点，以确定负责 4）测量PDPK的水平， 对照组和AD组脑的细胞质和颗粒组分。 两 将测量激酶活性和激酶蛋白水平。 这些研究 应该有助于澄清负责激酶的身份， PHF tau过度磷酸化。

The long term objective of this proposal is to understand the etiology and pathogenesis of Alzheimer disease (AD). Increasing evidence suggests that in AD the phosphorylation state of several neuronal proteins are altered. Such abnormal phosphorylation apparently lead to a disruption of different cellular functions and hence eventual death of neurons. In particular the microtubule-associated protein tau has been shown to be present in a hyperphosphorylated state in the paired helical filaments (PHF) of AD brain. Recent evidence suggests that the hyperphosphorylation of PHF tau may be catalyzed largely. but not exclusively, by proline-directed protein kinases (PDPKs). There are three well known PDPKs: MAP kinase, cdc2 kinase, and glycogen synthase kinase 3. The identities of the PDPKs and non-PDPKs responsible for the in vivo hyperphosphorylation of PHF tau are presently unclear. The specific aims of this proposal are: 1) To compare the phosphorylation of tau by the three PDPKs. The kinetics of phosphorylation and the sites phosphorylated on tau will be compared by peptide mapping and by different phosphorylation site-specific anti-tau antibodies; 2) To investigate the role of non-PDPKs in tau hyperphosphorylation. Six non- PDPKs will be used to prepare six species of phosphorylated tau. The further phosphorylation of these tau species by the PDPKs will be compared; 3) To use different synthetic peptides containing known phosphorylation sites of tau to establish the identities of the protein kinases responsible for their phosphorylation; and 4) To measure the levels of the PDPKs in both the cytosol and particulate fractions of control and AD brains. Both kinase activity and kinase protein levels will be measured. These studies should help clarify the identities of the kinase(s) responsible for the hyperphosphorylation of PHF tau.