FUNCTIONS OF ANKYRINS IN MICE

Ankyrins是一种大的、分布广泛的三体蛋白质，它连接着 整合的膜转运蛋白、黏附蛋白和受体 基于血影蛋白的膜骨架。已经克隆了三个Anyrin，并 到目前为止的特征：ANK1(红细胞锚蛋白)，Ank2(脑锚蛋白)， 和ANK3(上皮性锚蛋白)。每个锚链都含有一个N-末端 “重复”结构域将锚定蛋白连接到完整的膜蛋白上，一个 中央血影蛋白结合域，以及C-末端，“调节”域， 其功能没有被刻画出来。ANK3也存在于较小的 缺少重复结构域的异构体(ANK3-5kb)。巨噬细胞ANK3-5kb 仅与晚期溶酶体和溶酶体后相关 水泡。本研究旨在测试锚定蛋白的功能。 在体内，特别是ANK1是否可以被其他Anyrins取代 用ANK1替换原生ANK1会有什么后果 承受着监管领域的结构性变化。这些问题 将通过在小鼠身上进行靶向基因替换的技术来进行 胚胎干细胞。诱变对人骨肉瘤细胞的影响 生存、器官功能和病理、红细胞寿命和 形态、红细胞膜及膜骨架组成和 带的组织、膜的稳定性、功能和流动性 3人将接受调查。更多的实验将针对 识别以ANK3-5kb为靶点的细胞内小泡和 鉴定和克隆靶配体(S)。

Ankyrins are large, widely distributed, tripartite proteins that link integral membrane transporters, adhesive proteins and receptors to the spectrin-based membrane skeleton. Three ankyrins have been cloned and characterized so far: Ank1 (erythrocyte ankyrin), Ank2 (brain ankyrin), and Ank3 (epithelial ankyrin). Each ankyrin contains an N-terminal "repeat" domain that attaches ankyrin to integral membrane proteins, a central spectrin-binding domain, and a C-terminal, "regulatory" domain, whose function is not characterized. Ank3 also exists in a smaller isoform (Ank3-5kb)that lacks the repeat domain. In macrophages Ank3-5kb is associated exclusively with late lysosomes and post lysosomal vesicles. The present studies aim to test aspects of ankyrin function in vivo, particularly whether Ank1 can be replaced by the other ankyrins and what the consequences are of replacing native Ank1 with an Ank1 bearing structural changes in the regulatory domain. These questions will be pursued using the technique of targeted gene replacement in mouse embryonal stem (ES) cells. The effects of the induced mutations on survival, organ function and pathology, red cell life span and morphology, red cell membrane and membrane skeletal composition and organization, membrane stability, and the function and mobility of band 3 will be investigated. Additional experiments will be directed at identifying sequences that target Ank3-5 kb to intracellular vesicles and identifying and cloning the target ligand(s).