带状突触中RIBEYE蛋白介导的带状结构调控的分子机制研究

带状突触是一种特异性存在于感知性神经元如光感受器细胞、双极细胞、内耳细胞、松果体细胞等的突触结构。带状突触的突触前端活性区存在一个盘状或马蹄状的高电子密度致密区，其外周锚定了大量的突触小泡。 带状结构通过锚定和运输突触囊泡、招募钙离子通道等方式来调节神经递 质的释放，从而响应复杂的外界信息，比如声音、光、昼夜节律等信号范 围广、持续时间长、需要被及时响应的信号。RIBEYE 蛋白特异性存在于 带状突触中，是组成带状结构的主要成分。目前，对于 RIBEYE 蛋白如何 组装形成带状结构以及 RIBEYE 蛋白如何介导带状结构形态调节的过程等 问题还没有被研究清楚。为了回答这些问题，论文首先对 RIBEYE 蛋白进 行了序列保守性以及高级结构的分析。鉴定出 RIBEYE蛋白的保守区以及非保守区、折叠区和无序区，并将 RIBEYE 蛋白划分为不同的结构域，包 括：preSAM 结构域、SAM 结构域、 postSAM 结构域以及与 CtBP2 蛋白 高 度 同 源 的 B 结 构 域 。 通 过 在 HeLa 细 胞 中 过 表 达 RIBEYE 发 现 ， RIBEYE 蛋白在细胞中能通过相分离形成球状凝胶体。RIBEYE 蛋白通过 相分离可以形成两种形态的结构：球状结构、棒状结构。通过对不同结构 域的功能性分析，发现 RIBEYE 蛋白的 SAM 结构域、postSAM 结构域以 及 B 结构域对于棒状结构的形成至关重要。而且，RIBEYE 在细胞中的这 两种结构形态可以通过 preSAM 结构域进行调节。当存在 preSAM 时，棒 状结构被转换成球状结构。进一步分析发现，preSAM 结构域上的芳香族 氨基酸是调节这一转换的关键性氨基酸。当 preSAM结构域的芳香族氨基酸被突变后，preSAM 不再具有转换带状结构到球状结构的能力。综上所 述 ， 本 论 文 通 过 结 合 结 构 、 生 化 、 细 胞 等 多 种 实 验 分 析 方 法 ， 揭 示 了 RIBEYE 蛋白组装形成带状结构的关键性结构域，并进一步提出了调节带 状结构的可能性结构域以及关键性氨基酸。这些发现将有力地推动对带状 突触的功能和调控机制的理解。

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