Patterning mechanisms that restrict cardiac cushion formation in zebrafish

限制斑马鱼心脏垫形成的模式机制

• 批准号: 8716431
• 负责人: Andrew Robert Houk
• 金额: $ 2.62万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN DIEGO
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-05-01 至 2014-10-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8716431
• 关键词: Blood flow; Cardiac; Cells; Characteristics; Congenital Heart Defects; Data; Defect; Embryo; Endocardium; Equilibrium; Etiology; Event; Exposure to; Feedback; Future; Gene Expression; Genetic Epistasis; Goals; Heart; Heart Valves; Heat-Shock Response; Information Systems; Ketanserin; Knowledge; Laboratories; Life; Location; Molecular; Organism; Organogenesis; Pathway interactions; Pattern; Pharmaceutical Preparations; Phenotype; Play; Property; Reporter; Research; Research Personnel; Role; Serotonin; Serotonin Antagonists; Shapes; Signal Transduction; Structure; System; Techniques; Testing; Time; Transgenes; Tube; Ventricular; Zebrafish; cardiogenesis; cell type; congenital heart disorder; grasp; insight; malformation; mouse junctate protein; novel; optogenetics; public health relevance; receptor; research study; serotonin receptor; skills; small molecule; success; therapeutic target

DESCRIPTION (provided by applicant): Heart valves play a critical role in cardiac function by promoting unidirectional blood flow and are a frequent target of congenital heart disease. Atrioventricular valve formation is triggered by signals that are precisely localized to the junctin between chambers (also known as the atrioventricular canal or AVC). These signals instruct the local endocardium to bulge into the lumen of the heart tube, forming structures known as cardiac cushions that later remodel into valve leaflets. Studies in zebrafish have shown that canonical Wnt pathway activity at the AVC promotes the expression of genes encoding Bmp signals that induce cushion formation. Proper heart development necessitates the restriction of these inductive signals to the AVC, yet we do not understand the mechanisms responsible for this confinement. We also do not know important general properties of this system, such as whether Wnt signaling patterns itself or if it relays an upstream pattern. I will pursue two specifc aims to expand our knowledge of the components and systems-level properties of the AVC patterning network. My first aim investigates serotonin signaling, which our preliminary data suggest has a critical role in AVC patterning. Treatment of embryos with the serotonin receptor antagonist ketanserin results in ectopic bmp4 expression beyond the boundaries of the AVC. This phenotype is recapitulated by knockdown of the serotonin receptor htr2a, suggesting that serotonin signaling through this receptor is essential. I hypothesize that serotonin signaling participates in the confinement of cushion-inducing signals to the AVC. To test this hypothesis, I will first examine a panel of markers to thoroughly characterize the cushion patterning defect in htr2a-deficient embryos. I will next use pharmacological perturbations to determine the time window when serotonin signaling is critical for AVC patterning. I will also examine Wnt pathway readouts and perform epistasis experiments to determine whether serotonin acts upstream or downstream of the Wnt pathway to restrict cushion induction. Finally, I will use pharmacological perturbations of global serotonin levels in combination with htr2a misexpression to determine whether serotonin signaling is instructive or permissive for cushion restriction. In my second aim, I will test the hypothesis that Wnt signaling patterns itself, i.e. self- organizes. Self-organizing systems have several useful properties such as stability and robustness to fluctuations. These systems often contain a self-amplifying component (positive feedback) that is balanced by inhibition generated either by itself (autoinhibition) or by a rival self-amplifying component that dominates an adjacent domain (inhibitory crosstalk). I will use a combination of novel optogenetic techniques and live signaling reporters to determine whether Wnt signaling in the AVC displays hallmark characteristics of self-organizing systems, such as positive feedback, autoinhibition, and inhibitory crosstalk. Together, these studies will reveal important new mechanistic information and systems-level properties of the AVC patterning network, which will help us to understand the etiology of congenital heart defects.

描述（由申请人提供）：心脏瓣膜通过促进单向血流在心脏功能中发挥关键作用，是先天性心脏病的常见靶点。房室瓣的形成是由精确定位于房室交界处（也称为房室管或AVC）的信号触发的。这些信号指示局部内皮细胞膨胀到心管内腔中，形成称为心脏垫的结构，随后重塑为瓣膜小叶。在斑马鱼中的研究已经表明，AVC处的经典Wnt途径活性促进编码诱导垫形成的Bmp信号的基因的表达。正常的心脏发育需要限制这些诱导信号到AVC，但我们不了解这种限制的机制。我们也不知道这个系统的重要的一般属性，例如Wnt信号本身是否模式或它是否中继上游模式。我将追求两个具体的目标，以扩大我们的知识的组件和系统级属性的AVC图案网络。我的第一个目标是研究血清素信号，我们的初步数据表明，它在AVC模式中起着关键作用。用5-羟色胺受体拮抗剂酮色林处理胚胎导致异位bmp 4表达超出AVC的边界。这种表型通过5-羟色胺受体htr 2a的敲低而重现，表明通过该受体的5-羟色胺信号传导是必不可少的。我推测，5-羟色胺信号参与限制的cardion诱导信号的AVC。为了验证这一假设，我将首先检查一组标记物，以彻底表征htr 2a缺陷胚胎中的垫图案缺陷。接下来，我将使用药理学扰动来确定5-羟色胺信号传导对AVC模式至关重要的时间窗。我还将检查Wnt通路的读数，并进行上位性实验，以确定血清素是否作用于Wnt通路的上游或下游，以限制缓冲诱导。最后，我将结合htr 2a的错误表达，使用全身5-羟色胺水平的药理学扰动来确定5-羟色胺信号传导对缓冲限制是否具有指导性或许可性。在我的第二个目标中，我将测试Wnt信号本身模式的假设，即自组织。自组织系统具有几个有用的特性，例如稳定性和对波动的鲁棒性。这些系统通常包含一个自我放大的成分（正反馈），它被自身产生的抑制（自动抑制）或竞争对手的自我放大的抑制所平衡。 控制相邻域的分量（抑制性串扰）。我将使用新型光遗传学技术和活信号报告的组合来确定AVC中的Wnt信号传导是否显示自组织系统的标志性特征，如正反馈、自抑制和抑制性串扰。总之，这些研究将揭示AVC模式网络的重要新机制信息和系统级特性，这将有助于我们了解先天性心脏病的病因。