Nuclear Mechanics varies with Tissue Mechanics & Regulates Cytoskeleton

核力学随组织力学而变化

• 批准号: 8928873
• 负责人: Dennis E. Discher
• 金额: $ 23.33万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-09-14 至 2017-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8928873
• 关键词: Acute; Adult; Affect; Age of Onset; Aging; Animals; Aspirate substance; Automobile Driving; Back; Biomechanics; Birth; Blood; Blood Cells; Brain; Cardiac Myocytes; Cardiology; Cell Line; Cell Nucleus; Cell Survival; Cells; Cessation of life; Chick Embryo; Cicatrix; Collagen; Congenital Heart Defects; Connective Tissue; Cysteine; Cytoskeletal Gene; Cytoskeleton; Data; Development; Dilated Cardiomyopathy; Disease; Elasticity; Embryo; Embryonic Heart; Erythrocytes; Exhibits; Extracellular Matrix; Feedback; Feeds; Gel; Gene Expression; Genes; Genetic Transcription; Goals; Hair; Heart; Human; In Situ; Infarction; Keratin; Knock-out; Knockout Mice; Label; Lamin Type A; Lamin Type B; Lamins; Lead; Life; Link; Liquid substance; Marrow; Mathematics; Measurement; Measures; Mechanical Stress; Mechanics; Methods; Molecular; Molecular Conformation; Monitor; Mus; Mutation; Myosin Type II; Nail plate; Nuclear; Nuclear Lamin; Nuclear Lamina; Nuclear Structure; Organ; Pathway interactions; Peptides; Phosphorylation; Progeria; Proteins; Pump; Regulation; Relative (related person); Reporter; Retinoids; Rheology; Role; Science; Shotguns; Solid; Stem cells; Stress; Structural Protein; Structure; System; Testing; Therapeutic; Time; Tissue Differentiation; Tissues; Transcript; Transfection; Ursidae Family; Work; base; blastomere structure; blebbistatin; bone; embryo tissue; heart cell; mimetics; novel; promoter; public health relevance; regenerative; response; sensor; soft tissue; stoichiometry

 DESCRIPTION (provided by applicant): Biomechanical aspects of embryonic tissues are poorly understood, especially nuclear mechanics. Very early embryos are well-known to be very soft and have very low levels of the nuclear structure protein lamin-A, which we have shown by single cell manipulations means that the nucleus is softer than in almost any adult cell [Swift Science 2013]. Initial differentiation to tissue turns on lamin-A transcription in heart, which seems important because knockout mice exhibit 'developmental defects of the heart' and die shortly after birth [Kubben Nucleus 2011], but lamin-A protein characterization is lacking in intac embryonic tissues as studied here. Lamin-A mutations cause a range of diseases with various ages of onset, including dilated cardiomyopathy (DCM) and accelerated aging (Progeria) affecting heart. Lamin-A is also known to affect differentiation and cell survival - all of which motivates studies to see & perturb the lamina in beating hearts. With adult tissue and primary cells, we have found that lamin-A levels are nearly proportional to tissue stiffness E [Swift Science 2013]. Relatively stiff connective tissues bear high mechanical stress, such as bone and even heart, and they have high lamin-A, suggesting stiff nuclei resist the stress. In contrast, very soft tissues such as brain and marrow that bear little stress express low lamin-A. B-type lamins are comparatively constant in the solid tissues, so that lamin-A:B stoichiometry seems a mechanosensor of stiffness and stress in adult tissues. We have worked through the mathematics of a simple mechanobiological gene circuit that fits findings for adult cells and tissues. Our hypothesis here is that Lamins in normal embryos adjust developmentally in response to mechanical stresses. Our goal is to determine and perturb mechano- regulation of lamin gene circuits in developing embryos, with a focus on what develops into a stiff heart relative to fluid blood. We focus on the facile chick embryo system per our recent studies that demonstrate acute sensitivity of beating heart to matrix elasticity [Majkut Curr Biol 2013]. Chick has advantages including the fact that chick erythrocytes have lamins, but we will at the end compare to developing mouse tissues. First we will quantify lamin protein levels throughout development by Mass Spec, and we will assess their stress and stiffness sensitivity with novel measurements and perturbations. We will relate embryonic lamina measurements to nuclear rheology and perturb the levels to validate relationships and molecular mechanisms. Preliminary data shows that beating chick hearts are easily transfected, so that Lamin Promoter- Reporter constructs can be tested as in situ mechanosensors of stress and stiffness. The lamina also enhances maturation and differentiation, and initial data with adult cells indicates feedback to cytoskeletal gene expression and the retinoid pathway of therapeutic relevance. Our studies should ultimately reveal the nuclear lamina as a multi-factorial, embryonic stress sensor that feeds back into broader structural regulation. 16

 描述（由申请人提供）：人们对胚胎组织的生物力学方面知之甚少，尤其是核力学。众所周知，非常早期的胚胎非常柔软，并且核结构蛋白核纤层蛋白 A 的水平非常低，我们已通过以下方法证明了这一点。单细胞操作意味着细胞核比几乎所有成体细胞都要软 [Swift Science 2013] 组织的初始分化会启动心脏中的核纤层蛋白 A 转录，这似乎很重要，因为基因敲除小鼠表现出“发育性”。心脏缺陷”并在出生后不久死亡[Kubben Nucleus 2011]，但正如本文所研究的，完整胚胎组织中缺乏核纤层蛋白 A 突变，导致一系列不同年龄的疾病，包括扩张型心肌病。众所周知，核纤层蛋白 A (DCM) 和加速衰老 (早衰症) 也会影响心脏的分化和细胞存活——所有这些都促使人们研究和扰乱跳动心脏中的成体组织。和原代细胞中，我们发现核纤层蛋白-A 水平几乎与组织硬度 E 成正比 [Swift Science 2013] 相对坚硬的结缔组织承受高机械应力，例如骨骼甚至心脏，并且它们具有高核纤层蛋白-A，这表明。坚硬的原子核抵抗压力。 非常软的组织，如大脑和骨髓，承受的压力较低，在实体组织中表达的核纤层蛋白 B 型核纤层蛋白相对恒定，因此核纤层蛋白 A:B 的化学计量似乎是成人组织中硬度和压力的机械传感器。我们对一个简单的机械生物学基因回路进行了数学计算，该回路符合成人细胞和组织的发现。我们的假设是，正常胚胎中的层粘连蛋白会根据机械应力进行发育调整。扰乱发育中胚胎中核纤层蛋白基因回路的机械调节，重点关注相对于液体血液发展成僵硬心脏的因素。我们最近的研究表明，跳动的心脏对基质弹性非常敏感。 Majkut Curr Biol 2013]。小鸡具有优势，包括小鸡红细胞具有核纤层蛋白，但我们最后将与发育中的小鼠组织进行比较，首先我们将量化整个发育过程中的核纤层蛋白水平。通过质谱，我们将通过新的测量和扰动评估它们的压力和硬度敏感性，我们将把胚胎层测量与核流变学联系起来，并扰动水平以验证关系和分子机制，初步数据表明跳动的小鸡心脏很容易被转染。因此，核纤层蛋白启动子-报告基因结构可以作为应力和刚度的原位机械传感器进行测试。核纤层还可以增强成熟和分化，并且成体细胞的初始数据表明反馈。我们的研究最终应该揭示核纤层是一种多因素的胚胎应激传感器，可以反馈到更广泛的结构调节中。