RIN1, A NOVEL RAS-INHIBITORY PROTEIN IN YEAST

RIN1，酵母中一种新型 RAS 抑制蛋白

• 批准号: 6890919
• 负责人: DAVID E. LEVIN
• 金额: $ 25.34万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-06-01 至 2007-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6890919
• 关键词: binding sites; biological signal transduction; endoplasmic reticulum; fungal proteins; guanine nucleotide binding protein; membrane proteins; protein protein interaction; protein structure function; protein transport; site directed mutagenesis

DESCRIPTION (provided by applicant): The Ras family of small guanine 5'-triphosphatases (GTPases) comprise a group of molecular switches that link receptors on the cell surface to signaling pathways that regulate cell proliferation and differentiation. Ras proteins are highly conserved among eukaryotic species. Mutant forms of Ras that are locked in the guanine 5'-triphosphate (GTP)-bound state, and thus continuously drive cell proliferation by interacting with their target proteins, are found in 30 percent of human tumors. This application describes a novel protein in budding yeast that appears to serve two separate functions, despite the fact that it is only 68 amino acids in length. The gene encoding this protein, designated RIN1 (for Ras inhibitor 1) behaves genetically as an inhibitor of Ras, and the Rin1 protein associates in vivo with GTP-bound Ras in a manner that suggests it competes for the same binding site as Ras target proteins. Because Rin1 resides in the membrane of the ER, rather than on the cell surface with the majority of Ras, it is hypothesized that one function of Rin1 is to maintain Ras in an inactive state during its posttranslational modification in the ER. The long-term objective of this project is to develop a form of Rin1 that effectively inhibits GTP-bound Ras on the cell surface for the purpose of treating Ras-involved malignancies. The second function of Rin1 is in the processing or transport of glycosyl-phosphatidylinositol (GPI)-anchored proteins from the ER. It is hypothesized that this function is carried out through association with a second member of the Ras family that is involved in protein secretion. The specific aims of this project are: 1) To understand the membrane topology of Rin1, and to identify the Ras-interaction region. This is the first step toward developing a form of Rin1 that is optimized for Ras inhibition at the cell surface. 2) To understand the function of Rin1 in GPI-protein anchoring or secretion. This will involve identification of the second target of Rin1. It is also likely that this aim will contribute to the new area of protein sorting in the ER. 3) To determine if yeast Ras can signal from the ER, and to create a more effective Rin1-based inhibitor of Ras. This aim is focused on assessing the function of Ras expressed in the ER, and the function of the Ras-interacting domain of Rin1 expressed at the cell surface. These experiments will help to test the applicant's model of Rin1 function as an inhibitor of Ras signaling in the ER.

描述（由申请人提供）：小鸟嘌呤5 '-三磷酸酶（GTP酶）的Ras家族包含一组分子开关，其将细胞表面上的受体与调节细胞增殖和分化的信号传导途径连接。 Ras蛋白在真核生物中高度保守。 Ras的突变形式被锁定在鸟嘌呤5 '-三磷酸（GTP）结合状态，因此通过与其靶蛋白相互作用持续驱动细胞增殖，在30%的人类肿瘤中发现。 本申请描述了芽殖酵母中的一种新蛋白质，尽管其长度仅为68个氨基酸，但该蛋白质似乎具有两种独立的功能。 编码这种蛋白质的基因，命名为RIN 1（Ras抑制剂1），在遗传上表现为Ras的抑制剂，RIN 1蛋白在体内与GTP结合的Ras以一种方式结合，表明它与Ras靶蛋白竞争相同的结合位点。 由于Rin 1位于内质网的膜上，而不是与大部分Ras一起位于细胞表面，因此假设Rin 1的一个功能是在内质网中的翻译后修饰期间维持Ras处于非活性状态。 该项目的长期目标是开发一种形式的Rin 1，其有效抑制细胞表面上的GTP结合的Ras，以治疗Ras相关的恶性肿瘤。 Rin 1的第二个功能是从ER加工或转运糖基-磷脂酰肌醇（GPI）锚定蛋白。 据推测，这一功能是通过与Ras家族的第二个成员，参与蛋白质分泌的协会进行。 本项目的具体目标是：1）了解Rin 1的膜拓扑结构，并确定Ras相互作用区域。 这是开发Rin 1形式的第一步，该形式针对细胞表面的Ras抑制进行了优化。 2)了解Rin 1在GPI蛋白锚定或分泌中的作用。 这将涉及识别Rin 1的第二个靶标。 这一目标也可能有助于ER中蛋白质分选的新领域。 3)确定酵母Ras是否可以从ER发出信号，并创建更有效的Rin 1 Ras抑制剂。 本研究的目的是评估Ras在ER中表达的功能，以及Rin 1在细胞表面表达的Ras相互作用结构域的功能。 这些实验将有助于测试申请人的Rin 1作为ER中Ras信号传导抑制剂的功能模型。