MAINTENANCE OF LIPID ASYMMETRY IN THE HUMAN ERYTHROCYTE

描述：申请人注明其组成和组织 磷脂在人红细胞膜上的分布 对许多细胞的启动和调节至关重要 流程。而大多数与膜相关的酶和运输 功能与正常的膜脂不对称有关， 磷脂酰丝氨酸(PS)从其优势位置的再分布 在细胞的内部小叶到外部小叶导致一系列 似乎受其显示控制的事件。例如，ps 在细胞表面是止血的关键，并可能在 细胞老化、膜融合、识别和消除 衰老和凋亡的细胞。尽管已经取得了相当大的进展 为了解红血球中氨基磷脂的运动而做出的努力， 脂类运动的机制及其调节尚不清楚， 参与其控制或识别的蛋白质(S)也没有 已确认身份。 这项建议侧重于组织、动态和空间 PS在红细胞膜上的分布及其作用机制 网状内皮细胞识别老化的PS表达细胞。 特别强调的是识别和表征 调节PS分布和运动的蛋白质 红细胞中存在PS结合蛋白，巨噬细胞膜中存在PS结合蛋白。 拟议的研究是基于导致这一概念的结果 一种32 kDa的膜多肽，与Rh蛋白家族有关， 参与了膜脂不对称性的产生和调节。 应用程序的主要目标是隔离、识别和 PS结合蛋白及其相关蛋白的特性 跨双分子层脂质运动。这将通过以下组合来完成 技术包括从人工分离运输蛋白- 产生红细胞泡。巨噬细胞/单核细胞PS结合蛋白 将被PS亲和试剂分离出来。分离出的蛋白质将 进行测绘和测序。抗PS结合蛋白的抗体和 转运蛋白以及从信息中产生的寡核苷酸 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，以识别整个 转运蛋白和PS结合蛋白的编码区。结果是 这些研究将有助于理解监管 在体内维持特定跨双层脂质分布的过程 并介导人红细胞的识别和最终消除 衰老的PS表达细胞。

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.