MAINTENANCE OF LIPID ASYMMETRY IN THE HUMAN ERYTHROCYTE

人类红细胞中脂质不对称的维持

• 批准号: 2016336
• 负责人: ALAN Jay SCHROIT
• 金额: $ 17.71万
• 依托单位: UNIVERSITY OF TX MD ANDERSON CAN CTR
• 依托单位国家: 美国
• 财政年份: 1989
• 资助国家: 美国
• 起止时间: 1989-08-01 至 1999-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2016336
• 关键词: annexins; antibody; apoptosis; binding proteins; cell differentiation; cell senescence; erythrocyte membrane; erythrocytes; human tissue; laboratory mouse; lipid bilayer membrane; lipid biosynthesis; lipid transport; macrophage; membrane fusion; membrane reconstitution /synthesis; membrane structure; membrane transport proteins; monocyte; oligonucleotides; peptide library; phosphatidylserines; protein sequence; tissue /cell culture; vesicle /vacuole

DESCRIPTION: The applicants note that the composition and organization of phospholipids across the bilayer membrane of the human erythrocyte are critical to the initiation and regulation of many cellular processes. While most membrane-related enzymatic and transport functions are associated with normal membrane lipid asymmetry, the redistribution of phosphatidylserine (PS) from its preferential location in the cell's inner leaflet to the outer leaflet results in a sequence of events that appears to be regulated by its display. For example, PS at the cell surface is critical to hemostasis, and may play a role in cell aging, membrane fusion, and the recognition and elimination of senescent and apoptotic cells. Although considerable progress has been made toward understanding aminophospholipid movement in red blood cells, the mechanism of lipid movement and its regulation are not understood, nor have the protein(s) involved in its control or recognition been identified. This proposal focuses on the organization, dynamics and spatial distribution of PS in the erythrocyte membrane and the mechanism by which reticuloendothelial cells recognize aged, PS-expressing cells. Particular emphasis is placed on identifying and characterizing the proteins that regulate the distribution and control the movement of PS in erythrocytes and PS binding proteins present in macrophage membranes. The proposed studies are based on results that have led to the concept that a 32 kDa membrane polypeptide, related to the Rh family of proteins, is involved in the generation and regulation of membrane lipid asymmetry. The main objectives of the application are to isolate, identify, and characterize PS binding proteins and the proteins responsible for transbilayer lipid movement. This will be done by a combination of techniques including isolation of transport protein from artificially- generated erythrocyte vesicles. Macrophage/monocyte PS binding proteins will be isolated by PS affinity reagents. The isolated proteins will be mapped and sequenced. Antibodies against PS binding proteins and the transporter as well as oligonucleotides generated from information of the sequenced protein will be used to screen appropriate cDNA expression libraries. Positive clones will be sequenced to identify the entire coding region of the transporter and PS binding proteins. The results of these studies will contribute toward understanding the regulatory processes that maintain specific transbilayer lipid distributions in human erythrocytes and mediate the recognition and ultimate elimination of aged, PS-expressing cells.

说明：申请人注意到， 磷脂穿过人红细胞双层膜 对许多细胞的启动和调节至关重要 流程.虽然大多数与膜相关的酶和运输 功能与正常的膜脂不对称性有关， 磷脂酰丝氨酸（PS）从其优先位置重新分布 从细胞的内叶到外叶 似乎是由它的展示所调节的事件。例如，PS 在细胞表面的蛋白质对止血至关重要，并且可能在以下方面发挥作用： 细胞老化，膜融合，以及识别和消除 衰老和凋亡细胞。虽然取得了很大进展， 为了理解红细胞中的氨基磷脂运动， 脂质运动及其调节的机制尚不清楚， 参与其控制或识别的蛋白质也没有被 鉴定 该提案侧重于组织、动态和空间 PS在红细胞膜上的分布及其机制 网状内皮细胞识别衰老的表达PS的细胞。 特别强调的是， 调节PS分布和运动的蛋白质 在红细胞和PS结合蛋白存在于巨噬细胞膜。 拟议的研究是基于导致概念的结果 一种32 kDa的膜多肽，与Rh蛋白家族相关， 参与膜脂不对称性的产生和调节。 应用程序的主要目标是分离、识别和 表征PS结合蛋白和负责 跨双层脂质运动这将通过以下组合来完成： 技术包括从人工合成的蛋白中分离转运蛋白， 生成红细胞囊泡。巨噬细胞/单核细胞PS结合蛋白 将通过PS亲和试剂分离。分离的蛋白质将 绘制地图并排序。抗PS结合蛋白的抗体和 转运蛋白以及从转运蛋白的信息产生的寡核苷酸。 测序的蛋白质将用于筛选合适的cDNA表达 图书馆.将对阳性克隆进行测序，以确定整个 转运蛋白和PS结合蛋白的编码区。结果 这些研究将有助于了解监管 维持特定跨双层脂质分布的过程， 人红细胞并介导识别和最终消除 衰老的表达PS的细胞