Rho GTpase in the Lens - Role in Growth and Development

晶状体中的 Rho GTpase - 在生长和发育中的作用

• 批准号: 6719574
• 负责人: P VASANTHA Rao
• 金额: $ 26.95万
• 依托单位: DUKE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-03-01 至 2008-02-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6719574
• 关键词: DNA replication; JUN kinase; NAD(P)H dehydrogenase; apoptosis; biological signal transduction; cell differentiation; cell growth regulation; cell line; cell proliferation; clinical research; developmental neurobiology; enzyme activity; free radical oxygen; genetically modified animals; growth factor; guanine nucleotide binding protein; laboratory mouse; lens; mitogen activated protein kinase; nuclear factor kappa beta; organ culture; oxidative stress; protein tyrosine phosphatase; terminal nick end labeling; transfection /expression vector

DESCRIPTION (provided by applicant): The development of the ocular lens involves controlled proliferation and progressive differentiation of equatorial epithelial cells into terminally differentiated lens fibers. Intracellular signaling mechanisms that regulate these cellular processes and their possible role in cataractoqenesis have not been characterized. We hypothesize that Rho GTPases (Rho and Rac) play a critical role in lens growth and development and in maintenance of lens structural integrity by regulating cytoskeletal organization and growth factor stimulated stress-response signaling pathways. Strong evidence for this hypothesis derives from our recent studies based on pharmacological and genetic modulation of Rho GTPase function in different model systems including lens organ and epithelial cell culture, and a C3-exoenzyme transgenic mouse model. Inactivation of Rho GTPase in cell culture and in vivo resulted in alterations of lens epithelial and fiber cell morphology, actin cytoskeletal orqanization and cell adhesive characteristics. Additionally, the Rho GTPase functional knock-out mouse developed cataract and exhibited differential expression of genes encodinq proteins of the extracellular matrix and basement membrane, as well as proteins of stress signaling and apoptotic pathways. Further, several growth factors activated Rho GTPases (both Rho and Rac) in lens epithelial cells. These observations support a critical role for Rho GTPase (Rho and Rac)-mediated signaling mechanisms in lens development, growth and integrity. To elucidate the mechanistic basis (es) by which Rho GTPases regulate lens growth and development, we propose to investigate 1. The roles of RhoA and RhoB GTPases and their down stream effector-Rho kinase, in lens epithelial cell proliferation and cell fate using a RhoB knockout mouse, and by overexpressing dominant neqative mutants of RhoA or Rho kinase in a human lens epithelial cell line using adenoviral vectors. 2. The role of Rac GTPase and reactive oxygen species (ROS)-activated stress signaling pathways (C-jun-N-terminal kinase-JNK1 and p38 MAPK) in growth factor-induced proliferation and cell survival (Akt and NFkappaB) of human lens epithelial cells using an NADPH oxidase inhibitor and by overexpression of dominant negative and constitutively active mutants of Rac GTPase. 3. Regulation of lens protein tyrosine phosphatase activity by Rac/NADPH oxidase in organ-cultured lenses and in lens epithelial cells, in the context of lens growth and epithelial cell proliferation. The completion of these studies should unravel the significance of Rho/Rho kinase and Rac GTPase-mediated signaling and the role of stress -activated signaling pathways in proliferation, survival and apoptosis of lens epithelial cells. Better understanding of signal transduction pathways regulating lens growth and maintenance of lens transparency could provide insights into the etiology of cataract formation and into novel approaches towards developing medical treatments for certain types of cataract.

描述（由申请人提供）：眼镜的发育涉及赤道上皮细胞的受控增殖和逐步分化为末端分化的晶状体纤维。尚未表征调节这些细胞过程及其在白内障造成中的可能作用的细胞内信号传导机制。我们假设Rho GTPases（Rho和RAC）通过调节细胞骨架组织和生长因子刺激的应力 - 响应信号传导途径在晶状体生长和发展以及维持晶状体结构完整性方面起关键作用。该假设的有力证据源于我们最近的研究，基于不同模型系统中Rho GTPase功能的药理和遗传调节，包括镜头器官和上皮细胞培养，以及C3-示例性转基因小鼠模型。 Rho GTPase在细胞培养和体内的失活导致晶状体上皮和纤维细胞形态的改变，肌动蛋白细胞骨架orqanization和细胞粘合剂特征。此外，Rho GTPase功能基因敲除小鼠发展了白内障，并表现出细胞外基质和基础膜的基因的差异表达，以及应力信号传导和凋亡途径的蛋白质。此外，几个生长因子激活了晶状体上皮细胞中的Rho GTPase（Rho和RAC）。这些观察结果支持Rho GTPase（RHO和RAC）介导的信号传导机制在晶状体发育，生长和完整性中的关键作用。 To elucidate the mechanistic basis (es) by which Rho GTPases regulate lens growth and development, we propose to investigate 1. The roles of RhoA and RhoB GTPases and their down stream effector-Rho kinase, in lens epithelial cell proliferation and cell fate using a RhoB knockout mouse, and by overexpressing dominant neqative mutants of RhoA or Rho kinase in a human lens epithelial cell line使用腺病毒载体。 2。RAC GTPase和活性氧（ROS）激活的应力信号通路（C-Jun-N-末端激酶-JNK1和P38 MAPK）在生长因子诱导的增殖和细胞存活（AKT和NFKABPAB）中的增殖和细胞生存（AKT和NFKABPAB）中，使用人透镜上皮细胞和无性强化的强化量和纳经氧化剂的强制性强化， GTPase。 3。在晶状体生长和上皮细胞增殖的背景下，RAC/NADPH氧化酶在器官培养镜和晶状体上皮细胞中调节晶状体蛋白磷酸酶活性。这些研究的完成应阐明Rho/Rho激酶和RAC GTPase介导的信号传导的重要性，以及应激激活信号传导途径在晶状体上皮细胞增殖，存活和凋亡中的作用。更好地了解调节晶状体生长和透明度透明度维护的信号转导途径可以为白内障形成的病因提供见解，并进入为某些类型的白内障开发医疗治疗的新方法。