srGAP and the cadherin complex during morphogenesis in C. elegans

线虫形态发生过程中的 srGAP 和钙粘蛋白复合物

• 批准号: 9548360
• 负责人: Jeffrey D Hardin
• 金额: $ 11.7万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-08-01 至 2018-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9548360
• 关键词: Actins; Active Sites; Adherens Junction; Adhesions; Adhesiveness; Affect; Area; Binding; Biochemical; Biochemistry; Bundling; C-terminal; Cadherins; Caenorhabditis elegans; Cell membrane; Cell-Cell Adhesion; Cells; Complex; Congenital Abnormality; Cytoskeleton; Data; Defect; Dependence; Diagnosis; Embryo; Embryonic Development; Ensure; Epidermis; Epithelial; Epithelial Cells; Event; Genetic; Genetic Epistasis; Growth Cones; Human; Human Development; Image; In Vitro; Invaded; Lead; Maintenance; Maps; Mediating; Membrane; Mental Retardation; Modeling; Morphogenesis; Mutation; N-terminal; Neoplasm Metastasis; Organism; Orthologous Gene; Phospholipids; Point Mutation; Process; Property; Protein Family; Proteins; Proteomics; Recruitment Activity; Role; Signal Transduction; Site; Speed; Surface; System; Testing; Tissue Engineering; Tissues; Vertebrates; Work; base; cancer cell; cell behavior; functional genomics; human disease; in vivo; in vivo imaging; inducible gene expression; innovation; insight; migration; neuronal growth; novel; polymerization; public health relevance; tumorigenesis

DESCRIPTION (provided by applicant): Understanding what modulates the function of �-catenin, the key linker to the actin cytoskeleton within the cadherin-catenin complex (CCC) has widespread implications for understanding and treating defects during embryonic development, for modulating cell behavior in tissue engineering applications, and for diagnosing and treating metastatic tumors. Using functional genomics and proteomics, we identified the srGAP, SRGP-1, as an important regulator of the CCC during C. elegans morphogenesis. Mutations in srGAP orthologs lead to mental retardation in humans, but nothing is known about their roles in epithelial cell-cell adhesion. We will use genetics, innovative in vivo imaging, and biochemistry to test a model in which �-catenin and SRGP-1 act together to promote maturation and maintenance of adherens junctions in the following specific aims: Aim 1: Role of the SRGP-1 plasma membrane interaction in AJ maturation and maintenance. We will (a) characterize the phospholipid binding and bending properties of SRGP-1 using biochemistry, (b) assess recruitment of SRGP-1 to areas enriched in key phospholipids in vivo using dynamic imaging; and (c) test the importance of SRGP-1's membrane bending activity vs. general membrane recruitment in vivo using point mutations in key residues predicted to mediate convex vs. concave membrane bending and heterologous membrane targeting motifs. Aim 2: Role of the SRGP-1 interaction with actin regulators in AJ maturation and maintenance. We will (a) use a novel tissue-specific inducible expression system to test whether SRGP-1 regulates Rac signaling at nascent and maturing junctions in the epidermis; (b) use epitasis tests and WVE-1/deletion constructs predicted to abrogate SRGP-1 recruitment to the WAVE complex to place SRGP-1 within a genetic hierarchy during epidermal enclosure and embryonic elongation, and (c) we will test whether WVE-1/Wave, components of the C. elegans Sra-1/Nap1/Abi complex, and MIG-10/lamellipodin interact with the homologous region of SRGP-1. Aim 3: Role of SRGP-1 binding to �-catenin in AJ maturation and maintenance. We will (a) assess the temporal order of recruitment of SRGP-1 and the CCC using high-speed in vivo imaging, and test for their mutual dependence; (b) map the interacting domains within each protein, and assess the effects of the HMP-1/SRGP-1 interaction on their dynamics and function at AJs in vivo and on their known binding partners in vitro; and (c) we will validate candidate novel binding partners of the SRGP-1 C terminus, and perform large-scale screens for additional SRGP-1 physical interactors. As a result of these studies, we will gain new insight into how adherens junctions mature during epithelial morphogenesis in a living organism, a process crucial for diverse cellular events during human development and suppression of oncogenesis. We will also gain fundamental new insights in vivo into a class of proteins, the srGAPs, required for normal human development.

说明（应用程序提供）：了解哪种调节`catenin的功能，`catenin，catenin的关键接头是cadherin-catenin复合物（CCC）中肌动蛋白细胞骨架（CCC）在胚胎发育过程中的理解和治疗缺陷的范围，用于调节组织工程应用程序中的细胞行为，以便在组织工程应用中调节诊断和治疗METSTACTISTATICTACTITSTICTISTATICTACTISTATICTACTITSTICTISTATICTACTITSTICTISTACTITSTICTISTATICTACTITSTICTISTATICTACTITATTIS效果。使用功能性基因组学和蛋白质组学，我们将SRGAP SRGP-1确定为秀丽隐杆线虫形态发生过程中CCC的重要调节剂。 SRGAP直系同源物中的突变导致人类的智力低下，但对它们在上皮细胞 - 细胞粘合剂中的作用一无所知。我们将使用遗传学，创新的体内成像和生物化学来测试一个模型，在该模型中，`` - - - - - - - 钙蛋白和srgp-1）共同起作用，以促进以下特定目标的成熟和维持粘附连接的成熟和维护： 目标1：SRGP-1质膜相互作用在AJ成熟和维持中的作用。 我们将（a）使用生物化学来表征SRGP-1的磷脂结合和弯曲特性，（b）使用动态成像评估SRGP-1对体内关键磷脂中富集的区域的评估； （c）测试SRGP-1的膜弯曲活性与一般膜在体内募集的重要性，使用点突变在预测介质凸的关键保留率与凹形膜弯曲和靶向图基的异源膜的重要性。 目标2：SRGP-1与肌动蛋白调节剂的作用在AJ成熟和维持中。我们将（a）使用新型的组织特异性诱导表达系统来测试SRGP-1在表皮中是否调节RAC信号传导和成熟连接； （b）使用预测的表现表现测试和WVE-1/缺失构建体将招募SRGP-1募集到波复合物中，以将SRGP-1放置在表皮外壳和胚胎伸长期间的遗传层次结构范围内，并且（C）我们将与WVE-1/WAVE相互作用，无论是WVE-1/波是否与wve-1/abiip C. elemans sra-1/nap/nap1/abi and abip and abi and imig and nap1/abi and abiip and abi。 SRGP-1的同源区域。 AIM 3：SRGP-1在AJ成熟和维持中与 - 钙蛋白结合的作用。我们将（a）使用高速在体内成像中评估SRGP-1和CCC的临时募集顺序，并测试它们的相互依赖性； （b）绘制每种蛋白质中的相互作用结构域，并评估HMP-1/SRGP-1相互作用对其在体内AJS和其已知结合伴侣体外的动力学和功能的影响； （c）我们将验证SRGP-1 C末端的候选新型结合伙伴，并为其他SRGP-1物理相互作用器执行大规模筛选。由于这些研究，我们将获得新的见解 在生命组织中上皮形态发生过程中，粘附器连接如何成熟，这对于人类发育过程中的多元化细胞事件至关重要。我们还将在人体正常发展所需的一类蛋白质（SRGAP）中获得基本的新见解。