The F-BAR protein CIP4 in WAS-dependent thrombocytopenia

WAS依赖性血小板减少症中的F-BAR蛋白CIP4

• 批准号: 8322103
• 负责人: Seth Joel Corey
• 金额: $ 19.51万
• 依托单位: NORTHWESTERN UNIVERSITY AT CHICAGO
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-08-20 至 2014-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8322103
• 关键词: Actins; Address; Affect; Autoimmune Diseases; Autoimmune Process; Autoimmunity; B-Lymphocytes; Biology; Blood Platelet Disorders; Blood Platelets; Cardiovascular Diseases; Cardiovascular system; Cell membrane; Cell physiology; Cytoskeletal Modeling; Cytoskeletal Proteins; Defect; Disease; Dynamin; Eczema; Functional disorder; Genes; Genetic; Goals; Health; Hemorrhage; Human; Immune; Immunologic Deficiency Syndromes; Inflammation; Inflammatory; Knock-out; Knockout Mice; Label; Lead; Link; Lymphoma; Measures; Mediating; Megakaryocytes; Membrane; Metabolic Clearance Rate; Microtubules; Mouse Protein; Mouse Strains; Mus; Myelin P2 Protein; Natural Killer Cells; Neoplasm Metastasis; Pathway interactions; Phenotype; Phospholipids; Play; Production; Protein Binding; Protein Family; Proteins; Reporting; Risk; Role; SH3 Domains; Scaffolding Protein; Signal Pathway; Stroke; T-Lymphocyte; Testing; Thrombocytopenia; Vascular System; Wiskott-Aldrich Syndrome; Work; Yeasts; base; cdc42 GTP-Binding Protein; immune function; improved; insight; member; polymerization; protein function; rho GTP-Binding Proteins; src-Family Kinases; yeast two hybrid system

DESCRIPTION (provided by applicant): Platelets play a central role in cardiovascular, stroke, and bleeding disease through their quantity, activation, and interaction with the inflammatory and vascular systems. The Wiskott -Aldrich syndrome (WAS) is an X- linked disorder characterized by thrombocytopenia, immunodeficiency, eczema and an increased risk of lymphoma and autoimmunity. The affected gene encodes the cytoskeletal protein, WASP. Through the RhoGTPase, Cdc42, WASp undergoes conformational change that leads to actin polymerization. How WASp results in thrombocytopenia, the most common manifestation of WAS, remains unknown. Cdc42, Src kinases, membrane phospholipids, and SH3 domains control WASP function. Our lab has discovered a new regulator of WASP, CIP4 (Cdc42 interacting protein 4), in a yeast two hybrid screen with the Src kinase Lyn as bait. CIP4 is a member of the F-BAR family of proteins, which remodel the plasma membrane and promote actin polymerization. To determine CIP4's function, we created a CIP4-null mouse. We found that CIP4-/- mice display thrombocytopenia, similar to that observed in WASP-/- mice. The mechanism for thrombocytopenia in WAS is not known. Some studies demonstrate an autoimmune basis; others a defect in proplatelet production. Because our mice do not display signs of autoimmune disease, we favor the hypothesis ("defective proplatelet formation hypothesis") that CIP4-WASP forms a signaling pathway that promotes platelet production and that a deficiency of either CIP4 or WASP perturbs actin polymerization in megakaryocytes, which results in defective proplatelet formation. An alternative "immune destruction hypothesis" states that a defect in this CIP4-WASP pathway promotes autoimmunity and immune-mediated destruction of platelets. We propose to address these hypotheses through the following two specific aims: 1) the defective proplatelet hypothesis by culturing megakaryocyte precursors and megakaryocytes from wild-type, CIP4-/-, WASP-/-, and CIP4-/-WASP-/- mice and analyzing their proplatelet formation. We will use these unique mice strains to analyze proplatelet formation, the interaction of CIP4 with microtubules in megakaryocytes, and ultrastructural features of platelets; and 2) Test the immune destruction hypothesis by labeling platelets and measuring their clearance rates in wild-type, CIP4-/-, WASP-/-, and CIP4-/-WASP-/- mice and measuring their T, B, and NK cell functions. We will use these unique mice strains to measure platelet survival and to correlate with altered immune function.

描述（由申请人提供）：血小板通过其数量、激活以及与炎症和血管系统的相互作用在心血管、中风和出血性疾病中发挥核心作用。 Wiskott-Aldrich 综合征 (WAS) 是一种 X 连锁疾病，其特征为血小板减少、免疫缺陷、湿疹以及淋巴瘤和自身免疫风险增加。受影响的基因编码细胞骨架蛋白 WASP。通过 RhoGTPase、Cdc42，WASp 发生构象变化，导致肌动蛋白聚合。 WASp 如何导致血小板减少症（WAS 最常见的表现）仍不清楚。 Cdc42、Src 激酶、膜磷脂和 SH3 结构域控制 WASP 功能。我们的实验室在以 Src 激酶 Lyn 作为诱饵的酵母二杂交筛选中发现了 WASP 的新调节因子 CIP4（Cdc42 相互作用蛋白 4）。 CIP4 是 F-BAR 蛋白家族的成员，可重塑质膜并促进肌动蛋白聚合。为了确定 CIP4 的功能，我们创建了 CIP4-null 小鼠。我们发现 CIP4-/- 小鼠表现出血小板减少症，类似于在 WASP-/- 小鼠中观察到的情况。 WAS 血小板减少的机制尚不清楚。一些研究表明存在自身免疫基础；其他是前血小板产生的缺陷。因为我们的小鼠没有表现出自身免疫性疾病的迹象，所以我们支持这一假设（“缺陷性前血小板形成假说”），即 CIP4-WASP 形成促进血小板生成的信号通路，并且 CIP4 或 WASP 的缺陷会扰乱巨核细胞中的肌动蛋白聚合，这导致前血小板形成缺陷。另一种“免疫破坏假说”指出，CIP4-WASP 通路的缺陷会促进自身免疫和免疫介导的血小板破坏。我们建议通过以下两个具体目标来解决这些假设：1）通过培养来自野生型、CIP4-/-、WASP-/-和CIP4-/-WASP-/-小鼠的巨核细胞前体和巨核细胞来提出有缺陷的前血小板假说并分析它们的前血小板形成。我们将使用这些独特的小鼠品系来分析前血小板的形成、CIP4与巨核细胞中微管的相互作用以及血小板的超微结构特征； 2) 通过标记血小板并测量其在野生型、CIP4-/-、WASP-/- 和 CIP4-/-WASP-/- 小鼠中的清除率并测量其 T、B 和 NK 来测试免疫破坏假说细胞功能。我们将使用这些独特的小鼠品系来测量血小板存活率并与改变的免疫功能相关联。