Differential binding of apolipoprotein E isoforms to tau and other cytoskeletal proteins
Differential binding of apolipoprotein E isoforms to tau and other cytoskeletal proteins
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DOI：
10.1006/exnr.1996.0064
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发表时间：
1996-04-01
影响因子：
5.3
通讯作者：
Johnson, GVW
中科院分区：
文献类型：
--
作者：
Fleming, LM;Weisgraber, KH;Johnson, GVW
载脂蛋白 E4 (apoE4) 基因剂量是迟发性和散发性阿尔茨海默病的主要危险因素，50% 的纯合子患者在 70 岁时患上这种疾病。之前的研究表明，apoE 定位于大脑内某些神经元的细胞质。此外，apoE3（而非 apoE4）与微管相关蛋白 tau 和 MAP-2 形成 SDS 稳定的复合物。为了扩展这些研究并定量 apoE 与其他蛋白质的关联，使用凝胶位移和叠加测定检查了 apoE3 和 apoE4 与几种细胞骨架蛋白的关联。在凝胶迁移测定中，apoE3 与人重组 tau 最长同工型 (T4L)、人重组 tau 最短同工型 (T3) 和 160 kDa 神经丝蛋白 (NFM) 形成 SDS 稳定复合物，在该测定中 ApoE4 不结合 T3、T4L 或 NFM。在重叠测定中进一步检查apoE3和apoE4与T4L、肌动蛋白或微管蛋白的关联，将已知量的细胞骨架蛋白印迹到硝酸纤维素上并在0.15μM（5μg/ml）apoE3或apoE4中孵育。在此测定中，apoE3和apoE4同样良好地结合T4L和微管蛋白。相反，apoE3以比apoE4显着更高的亲和力结合肌动蛋白。这些结果表明，apoE 亚型与细胞骨架蛋白以至少两种不同的结合亲和力相互作用。更强烈的相互作用导致形成 SDS 稳定的复合物，并且几乎仅与 apoE3 发生，而 apoE 和细胞骨架蛋白之间的其他相互作用对于 apoE3 并不具有特异性。 (C) 1996 学术出版社
The apolipoprotein E4 (apoE4) gene dose is a major risk factor for late-onset and sporadic Alzheimer's disease with 50% of homozygous patients developing the disease by age 70. Previous studies have shown localization of apoE to the cytoplasm of certain neurons within the brain. In addition, apoE3, but not apoE4, forms SDS-stable complexes with the microtubule-associated proteins tau and MAP-2. To extend these studies and quantitate the association of apoE with other proteins, the association of apoE3 and apoE4 with several cytoskeletal proteins was examined using both gel shift and overlay assays. In the gel shift assay, apoE3 formed SDS-stable complexes with the longest isoform of human recombinant tau (T4L), the shortest isoform of human recombinant tau (T3), and the 160-kDa neurofilament protein (NFM), ApoE4 did not bind T3, T4L, or NFM in this assay. The association of apoE3 and apoE4 with T4L, actin, or tubulin was further examined in an overlay assay with known amounts of the cytoskeletal proteins slot-blotted onto nitrocellulose and incubated in 0.15 mu M (5 mu g/ml) apoE3 or apoE4. In this assay, apoE3 and apoE4 bound T4L and tubulin equally well, In contrast, apoE3 bound actin with a significantly greater affinity than did apoE4. These results indicate that apoE isoforms interact with cytoskeletal proteins with at least two different binding affinities. The more avid interaction results in the formation of complexes which are SDS stable and occurs almost exclusively with apoE3, while the other interactions between apoE and cytoskeletal proteins are not specific for apoE3. (C) 1996 Academic Press, Inc.