Mechanisms underlying asymmetric rotation and morphogenesis of the midgut

中肠不对称旋转和形态发生的机制

• 批准号: 10522575
• 负责人: Natasza A Kurpios
• 金额: $ 40万
• 依托单位: CORNELL UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-04-01 至 2025-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10522575
• 关键词: Actomyosin; Address; Affect; Alleles; Anisotropy; Architecture; BMP4; Bilateral; Birds; Blood Vessels; Cell Polarity; Cell physiology; Cells; Chick Embryo; Chickens; Computer Models; Congenital Abnormality; Cytoskeleton; Data; Development; Diagnosis; Diagnostic; Disease; Dorsal; Dose; Embryology; Embryonic Development; Endothelial Cells; Enzymes; Equilibrium; Extracellular Matrix; Failure; Feedback; Gene Expression; Gene Targeting; Genes; Genetic; Genetic Transcription; Goals; Handedness; Hyaluronan; Intestinal Volvulus; Intestines; Isomerism; Knockout Mice; Left; Life; Link; Live Birth; Location; Lymphatic; Lymphedema; Malignant Neoplasms; Measurement; Mechanics; Mesenchymal; Mesenchyme; Mesentery; Midgut; Modeling; Molecular; Morphogenesis; Mus; Muscle; Myosin ATPase; Newborn Infant; Nodal; Organ; Organogenesis; Pattern; Physical condensation; Process; Property; Regulation; Rieger syndrome; Role; Rotation; Shapes; Side; Signal Transduction; Stress; Suggestion; System; Technology; Testing; Time; Tissues; Translating; Translations; Tube; Vascular blood supply; Vascularization; Vertebrates; Work; cell behavior; cell motility; chicken egg; clinically significant; craniofacial; dosage; driving force; fascinate; gain of function; genetic manipulation; improved; lymphatic development; lymphatic malformations; lymphatic vessel; mechanical force; mechanotransduction; mouse genetics; mouse model; mutant; neonate; pediatric patients; prevent; quantitative imaging; response; tool; vasculogenesis

ABSTRACT The mechanisms discovered through the study of embryogenesis have been fundamental to understanding disease. We use classic chicken embryology and sophisticated mouse genetics to elucidate how basic cellular processes define the shape and function of organs. We are most fascinated by left-right (LR) organ asymmetry, as errors of organ laterality are linked to life-threatening birth defects. The counterclockwise rotation of the gut is an excellent model to study organ laterality. A critical aspect of this rotation is initiation of a leftward tilt directed by the master regulator of LR asymmetry, Pitx2. Failure to do so leads to gut malrotation and catastrophic volvulus in pediatric patients. Whereas rotation forces had long been assumed intrinsic to the gut tube, we instead discovered that gut rotation is driven by asymmetric deformation of the adjacent dorsal mesentery (DM) that suspends the gut, and whose cellular architecture is downstream of Pitx2. A key property of the DM is its exquisite binary organization, with distinct LR compartments that are readily accessible to genetic manipulation. Cellular and extracellular matrix (ECM) changes in each compartment cause the DM to deform and tilt the attached gut tube leftward. This critical bias determines gut chirality and frames a model to explain how LR gene expression is ultimately responsible for changes in cell behavior that initiate asymmetric organogenesis. The DM is also the sole conduit for blood and lymphatic vessels that serve the gut. We discovered that Pitx2-dependent mechanisms directing gut tilting are also crucial for patterning the gut vasculature and provide a mechanism to coordinate these two processes. Whereas most situs-specific organogenesis depends on Pitx2, mechanistic studies downstream have been hampered by a confounding “double-right” isomerism in Pitx2 mutants. Whereas Pitx2 expression in all vertebrates is activated by Nodal, Nodal disappears before asymmetric morphogenesis, leaving unresolved the question of how Pitx2 directs organogenesis. We discovered that Pitx2 expression in the gut is not an extension of previous induction by Nodal. Instead, we demonstrate that gut rotation requires a “second wave” of Pitx2 that is subject to mechanoregulation by the latent TGFb, linking LR gene expression to force translation. In aim 1, we determine the mechanism of Pitx2 dose response during gut, vascular, and lymphatic development and identify two distinct roles for Pitx2 dependent on its repressive threshold on BMP4 signaling. In aim 2, we define the molecular mechanism by which the formin Daam2, a Pitx2 target, directs mesenchymal cell polarity, actomyosin contractile asymmetry, and thereby steers the forces to polarize tilting. In aim 3, we combine cutting-edge physical measurements of tissue properties with quantitative imaging and computational modeling, to elucidate how stiffness and force anisotropies drive gut rotation through mechanical feedback. Together, these studies will significantly advance our understanding of the transcriptional and mechanical control of asymmetric gut and vascular morphogenesis, a critical step toward improved malrotation diagnostics in newborns.

摘要 通过胚胎发生研究发现的机制一直是理解 疾病。我们使用经典的鸡胚胎学和复杂的小鼠遗传学来阐明基本的细胞 过程定义了器官的形状和功能。我们最着迷的是左右器官的不对称， 因为器官偏侧的错误与危及生命的出生缺陷有关。肠道的逆时针旋转 是研究器官偏侧性的极佳模型。这种旋转的一个关键方面是开始向左倾斜。 由LR不对称的主调节器PITX2指示。如果不这样做，会导致肠道旋转不良和 儿科患者的灾难性扭转。而旋转力长期以来一直被认为是肠道的固有力量 管，相反，我们发现肠道旋转是由邻近背部的不对称变形驱动的 肠系膜(DM)，悬挂肠道，其细胞结构位于PITX2下游。一个重要的财产 DM的特点是其精致的二进制组织，具有独特的LR隔间，易于访问 基因操纵。细胞和细胞外基质(ECM)在每个隔室的变化导致DM 将连接的肠管向左变形和倾斜。这一关键偏差决定了肠道手性，并将模型构建为 解释LR基因的表达如何最终导致引发不对称的细胞行为变化 器官发生。糖尿病也是供应肠道的血液和淋巴管的唯一管道。我们 发现依赖Pitx2的机制引导肠道倾斜对肠道模式也是至关重要的 血管系统，并提供一种机制来协调这两个过程。而大多数站点特定的 器官发生依赖于PITX2，下游的机制研究因一种令人困惑的 Pitx2突变体的“双右”异构性。虽然所有脊椎动物中的Pitx2表达都是由Nodal激活的， 结节在不对称形态发生之前就消失了，留下了一个悬而未决的问题，即Pitx2如何 引导器官发生。我们发现，Pitx2在肠道中的表达不是先前诱导的延伸 由诺达尔。相反，我们证明肠道旋转需要Pitx2的“第二波”，它受制于 通过潜在的TGFb进行机械调节，将LR基因的表达与强制翻译联系在一起。在目标1中，我们确定 肠、血管和淋巴发育过程中Pitx2的剂量反应机制及鉴定 Pitx2的不同作用取决于其对BMP4信号的抑制阈值。在目标2中，我们定义了 作为PITX2靶标的Forin Daam2引导间充质细胞极性肌动球蛋白的分子机制 收缩不对称，从而引导力量极化倾斜。在目标3中，我们结合了尖端的物理 用定量成像和计算模型测量组织特性，以阐明如何 刚度和力的各向异性通过机械反馈驱动肠道旋转。总而言之，这些研究将 极大地促进了我们对不对称肠道的转录和机械控制的理解 血管形态发生，改善新生儿旋转不良诊断的关键一步。