Membrane skeleton regulation of cell shape and interactions in lens development

细胞形状的膜骨架调节和晶状体发育中的相互作用

• 批准号: 7680014
• 负责人: Velia M Fowler
• 金额: $ 47.38万
• 依托单位: SCRIPPS RESEARCH INSTITUTE, THE
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-09-01 至 2012-08-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7680014
• 关键词: Actins; Adult; Age; Alleles; Anterior; Architecture; Binding; Biochemical; Biological Assay; Cataract; Cell Differentiation process; Cell Maturation; Cell Shape; Cell membrane; Cell-Cell Adhesion; Cells; Cellular Structures; Chickens; Collaborations; Complex; Crystalline Lens; Crystallins; Data; Defect; Development; Disease; Dissociation; Embryo; Employee Strikes; Erythrocytes; Family; Filament; Goals; Growth; Growth and Development function; Heart; Human; Individual; Inherited; Knock-out; Knockout Mice; Laboratories; Lasers; Lead; Length; Lens Fiber; Life; Maintenance; Measures; Mechanics; Membrane; Microfilaments; Minus End of the Actin Filament; Molecular; Morphogenesis; Mus; Mutation; Myosin Heavy Chains; Optics; Pattern; Phenotype; Precipitation; Progress Reports; Property; Protein Isoforms; Proteins; Proteolysis; Proteolytic Processing; Protocols documentation; Regulation; Regulation of Cell Shape; Research; Role; Scanning; Shapes; Spectrin; Structure; Testing; Tissues; Transgenes; Transgenic Mice; Tropomyosin; Work; adhesion receptor; age related; base; depolymerization; design; fiber cell; genetic regulatory protein; in vivo; insight; lens; lens cortex; lens morphogenesis; lens protein; lens transparency; light scattering; membrane skeleton; migration; monomer; mouse model; mutant; postnatal; programs; promoter; protein function; public health relevance; response

DESCRIPTION (provided by applicant): The ocular lens is comprised of successive layers of extremely long thin fiber cells whose shapes, ordered hexagonal packing and regular membrane structure are critical for lens transparency and focusing. The broad, long-term objectives of this research are to elucidate the role of the spectrin-based membrane skeleton in fiber cell morphogenesis, lens optical quality and mechanical properties, and how defects in the membrane skeleton may contribute to progressive, age-dependent declines in lens functions. This proposal will study tropomodulins (Tmods), a family of actin regulatory proteins that function cooperatively with tropomyosins (TMs) to cap actin filament pointed ends, regulating actin filament turnover and stability in the membrane skeleton, contributing to cell shapes, interactions and mechanical properties. The mouse lens contains two Tmods, Tmod1 and Tmod3, which are both associated with fiber cell membranes. To investigate Tmod functions in vivo in the lens, we have generated knockout mice with targeted deletions in Tmod1 and Tmod3. Recent work from our laboratory has shown that lenses lacking Tmod1 develop normally but demonstrate striking defects in their cortical fiber cells, including abnormal fiber cell shapes with disordered cellular packing in the anterior cortex. Levels of a membrane-associated, short 3-TM isoform are reduced selectively in the absence of Tmod1, concomitant with depolymerization of actin filaments and reduction of 12-spectrin on membranes. It is hypothesized that Tmod1 and the short 3-TM function to stabilize the actin filament linkers in the membrane skeleton, thus controlling its integrity and long-range organization. This is expected to be critical for anchoring of adhesion receptors to control fiber cell shapes, interactions and ordered packing. Increased levels of Tmod3 in embryonic and postnatal but not adult lenses indicates that Tmod3 may compensate for the absence of Tmod1 in lens development and initial growth, leading to age-dependent progression of fiber cell disorder. Further, Tmod3 (but not Tmod1) is proteolysed in adult lens fiber cells, suggesting a unique role for Tmod3 degradation in membrane skeleton remodeling in fiber cell elongation and maturation. The specific aims are: (1) To investigate the role of Tmod1 in controlling membrane skeleton assembly, stability and associations with adhesion receptors by biochemical and morphological analyses of lenses from Tmod1 knockout mice, (2) To investigate the expression and function of Tmod3 in lens fiber cells and its role in compensating for absence of Tmod1 by molecular, biochemical and morphological analyses of lens development and growth in Tmod1 and Tmod3 knockout mice, (3) To investigate the consequences of loss of Tmod1 or Tmod3 for lens optical quality and mechanical stiffness by functional assays on living lenses ex vivo. Completion of these studies will elucidate the molecular and cellular basis for membrane skeleton regulation of fiber cell shapes and interactions during lens development, growth, and ageing, and how this influences lens optical quality and mechanical properties. PUBLIC HEALTH RELEVANCE: This project seeks to elucidate the molecular basis for membrane skeleton regulation of lens fiber cell shape, interactions and ordered hexagonal packing, and how this contributes to maintenance of lens transparency and focusing ability. Our studies will utilize transgenic mouse models in which deficiencies in specific membrane skeleton components lead to developmental or age-dependent abnormalities in lens fiber cell structures and interactions. These studies are expected to provide mechanistic insights into the molecular and cellular basis for normal lens optical quality and mechanical functions. Our studies on mouse lenses may also aid in understanding the causes of inherited human cataracts and loss of accommodative ability with age.

描述（由申请人提供）：眼镜由极长的薄纤维细胞的连续层组成，其形状，有序的六边形堆积和常规膜结构对于透镜透明度和聚焦至关重要。这项研究的广泛，长期的目标是阐明基于光谱的膜骨架在纤维细胞形态发生，晶状体光学质量和机械性能以及膜骨架中的缺陷中如何有助于晶状体功能的渐进，年龄依赖性下降。该提案将研究肌动蛋白的肌动蛋白调节蛋白家族（TMODS），该蛋白质与肌动蛋白（TMS）合作起作用，以粘贴肌动蛋白丝尖的末端，调节肌动蛋白丝的转移和膜骨架中的稳定性，从而有助于细胞形状，相互作用，相互作用和机械性能。小鼠透镜包含两个TMODS TMOD1和TMOD3，它们都与纤维细胞膜相关。为了研究晶状体中体内的TMOD功能，我们在TMOD1和TMOD3中产生了具有靶向缺失的基因敲除小鼠。我们实验室的最新工作表明，缺乏TMOD1的镜头正常发展，但表现出其皮质纤维细胞中的醒目缺陷，包括异常的纤维细胞形状，并在前皮层中具有无序的细胞堆积。在没有TMOD1的情况下，选择性降低膜相关的短3-TM同工型水平，与肌动蛋白丝的解聚并减少膜上12-谱蛋白。假设TMOD1和简短的3-TM功能可以稳定膜骨架中的肌动蛋白丝接头，从而控制其完整性和远距离组织。预计这对于锚定粘附受体控制纤维细胞形状，相互作用和有序填料至关重要。胚胎和产后但不是成年镜头中TMOD3水平的升高表明，TMOD3可能会弥补晶状体发育和初始生长中TMOD1的缺失，从而导致纤维细胞疾病的年龄依赖性进展。此外，TMOD3（而非TMOD1）是在成年晶状体纤维细胞中蛋白溶质化的，这表明TMOD3降解在膜骨架重塑中在纤维细胞伸长和成熟中起着独特的作用。具体目的是：（1）调查TMOD1在控制膜骨骼组装，稳定性以及与粘附受体中的作用，通过生化和形态学分析TMOD1敲除小鼠的透镜（2），（2）研究TMOD3在晶状体纤维细胞中的表达和功能，以补偿TMOD，以补偿TMOD的作用。 TMOD1和TMOD3基因敲除小鼠的晶状体发育和生长的分析，（3）研究了TMOD1或TMOD3损失对透镜的光学质量和机械刚度的损失，这是通过对生物镜头的功能分析的。这些研究的完成将阐明晶状体发育，生长和衰老过程中纤维细胞形状和相互作用的膜骨架调节的分子和细胞基础，以及这如何影响透镜的光学质量和机械性能。公共卫生相关性：该项目旨在阐明透镜纤维细胞形状，相互作用和有序六角形堆积的膜骨架调节的分子基础，以及这如何有助于维持镜片透明度和聚焦能力。我们的研究将利用转基因小鼠模型，其中特定膜骨骼成分的缺陷导致透镜纤维细胞结构和相互作用的发育或年龄依赖性异常。这些研究有望为正常晶状体光学质量和机械功能提供对分子和细胞基础的机械见解。我们对小鼠透镜的研究也可能有助于了解遗传性人白内障的原因以及随着年龄的增长而失去了适应性的能力。