MECHANISM OF SEGREGATION OF MODIFIED TUBULIN IN VIVO

微管（MT）是细胞骨架元素，存在于所有细胞中。 真核细胞，在那里它们在各种运动和 形态发生事件 所有MT都是微管蛋白的聚合物 原聚体是一种长寿命的细胞蛋白质， 在不同的生物体和细胞类型中是非常恒定的。 这项研究的长期目标是了解 微管蛋白或其相关蛋白质的生化变化 可以改变MT的功能。 特别是， 转录后修饰似乎是好的， 可能会迅速而可逆地影响 MT组织或功能。 微管蛋白受一对独特的 翻译后修饰，称为 酪氨酸化/去酪氨酸化，其中加入酪氨酸残基 连接到α链的C末端或从α链的C末端去除。 我们 以前发现MT富含酪氨酸或酪氨酸磷酸化， 去酪氨酸微管蛋白组成细胞的互补亚群， MT，并且这些微管蛋白种类通过环状微管蛋白分离。 涉及酪氨酸化/去酪氨酸化和微管蛋白的机制 聚合/解聚。 我们还证明 富含脱酪氨酸微管蛋白的MT表现出不寻常的 性质和动力学。 在这里，我们建议对此进行进一步的研究。 不寻常的稳定、脱酪氨酸MT人群，以及 确定这些MT的哪些属性是以下各项的函数： 脱酪氨酸原聚体的存在。 此外，我们将 对酪氨酸酶（微管蛋白）进行详细的研究 酪氨酸连接酶）和脱酪氨酸（微管蛋白羧肽酶）α- 微管蛋白;纯化每一个，测定其活性和MT过程中的水平， 介导的细胞事件，衍生每个的cDNA克隆， 将其特性与其他相关酶进行比较。 最后，酶和不寻常的MT的表征 他们所创造的将使我们能够开发化学和反义RNA， 每种酶的抑制剂，以确定的作用， MT功能中的酪氨酸/脱酪氨酸周期。 我们提出的 对这种独特的翻译后修饰循环的研究将 使我们能够更好地了解MT在 分化和细胞分裂在正常和病理 设置.

Microtubules (MTs) are cytoskeletal elements found in all eukaryotic cells, where they function in a variety of motile and morphogenetic events. All MTs are polymers of tubulin protomers, which are long-lived cellular proteins whose sequence is remarkably constant among different organisms and cell types. The long-range goal of the research proposed is to understand what biochemical alterations in tubulin or its associated proteins can bring about changes in the function of MTs. In particular, post-transcriptional modifications would seem to be good candidates for alterations that could rapidly and reversibly affect MT organization or function. Tubulin is subject to a unique pair of post-translational modifications, called tyrosination/detyrosination, in which a tyrosine residue is added to or removed from the C-terminus of the alpha chain. We previously found that MTs enriched in tyrosinated or detyrosinated tubulin composed complementary subsets of cellular MTs, and that these tubulin species become segregated by a cyclic mechanism involving tyrosination/detyrosination and tubulin polymerization/depolymerization. We have also demonstrated that MTs enriched in detyrosinated tubulin exhibit unusual properties and dynamics. Here we propose further studies of this unusual population of stable, detyrosinated MTs, and a determination of which properties of these MTs are a function of the presence of detyrosinated protomers. In addition, we will perform detailed studies on the enzymes that tyrosinate (tubulin tyrosine ligase) and detyrosinate (tubulin carboxypeptidase) alpha- tubulin; purifying each, assaying its activity and levels during MT- mediated cellular events, deriving cDNA clones to each and comparing its properties to those of other related enzymes. Finally, characterization of the enzymes and the unusual MTs they create will allow us to develop chemical and anti-sense RNA inhibitors of each enzyme, in order to determine the role of the tyrosination/detyrosination cycle in MT function. Our proposed work on this unique cycle of post-translational modification will enable us to better understand the function of MTs during differentiation and cell division in both normal and pathological settings.