Identifying long-range polarity cues using spontaneous mutations in mammals

利用哺乳动物的自发突变识别长程极性线索

• 批准号: 10454297
• 负责人: Maureen Patricia Cetera
• 金额: $ 24.9万
• 依托单位: UNIVERSITY OF MINNESOTA
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-07-20 至 2024-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10454297
• 关键词: Affect; Animals; Architecture; Artificial Organs; Back; Behavior; Biological Models; Cavia; Cell Communication; Cells; Cellular biology; Cilia; Complex; Congenital Heart Defects; Cues; Data; Defect; Development; Developmental Biology; Disease; Drosophila genus; Embryo; Embryonic Development; Ensure; Epidermis; Etiology; Failure; Feedback; Generations; Genes; Genetic; Genetic Screening; Genomics; Goals; Growth; Growth and Development function; Hair follicle structure; Head; Image Analysis; In Vitro; Integral Membrane Protein; Labyrinth; Lesion; Light; Live Birth; Mammals; Methods; Molecular; Morphogenesis; Mus; Mutant Strains Mice; Mutation; Nature; Neural Tube Closure; Organ; Pathway interactions; Pattern; Phenotype; Prize; Process; Proteins; Research; Side; Signal Pathway; Signal Transduction; Skin; System; Tissue Expansion; Tissues; Work; causal variant; congenital heart disorder; developmental disease; extracellular; gene product; genetic analysis; hearing impairment; in vivo; malformation; mutant; novel; planar cell polarity; prenatal therapy

Project Summary/ Abstract Neural tube closure defects and congenital heart defects are common developmental disorders, affecting 1 in 1000 and 1 in 100 live births, respectively. During embryonic development, cells must be able to sense direction and coordinate collective behaviors to avoid these severe malformations. Although the signaling pathway that imparts directionality between neighboring cells is known, uncovering the mechanisms that transmit this information on the order of thousands of cells has stalled. Fortuitously, spontaneous mutations in small mammals have given rise to animals that are prized for region specific disruptions in fur orientation. I hypothesize that the mutations disrupt long-range directional cues. The objective of this proposal is to utilize spontaneous mutations to identify long-range directional cues in the mammalian epidermis. This work will shed light on the etiology of complex congenital diseases, which will assist in the development of in utero therapies and the generation of artificial organs. A key molecular pathway that allows cells to communicate directional information is the Planar Cell Polarity (PCP) pathway, without which severe developmental defects arise. The core pathway consists of three transmembrane proteins that form asymmetric complexes that are localized to opposite sides of a cell. Through both positive extracellular feedback and negative intracellular feedback, this asymmetry is propagated locally but the mechanism that orients asymmetry across entire tissues is poorly understood. Although the core pathway components were identified through forward genetic screens in Drosophila, this method has been less successful in uncovering global polarity cues. The mouse skin provides an ideal system to identify long-range polarity cues. The expansive tissue is decorated with thousands of hair follicles (HFs) that are all oriented in the same direction through a PCP driven process. Remarkably, we showed the back skin is compartmentalized into regional domains that influence the direction of PCP signaling. This data suggests that region specific long-range cues cooperate to coordinate PCP and HF orientation across the entire tissue. In support of this idea, spontaneous mutations in small mammals produce animals with distinct, region-specific disruptions in hair follicle orientation. To investigate whether these mutations affect long-range polarity genes, I will identify the causative mutations through genomic sequencing and characterize the effect the mutations have on tissue polarity using genetics and image analysis. I will then interrogate the ability of the identified proteins to act as long-range polarity cues in vivo and in organotypic skin culture. Upon completion of this study, we will identify and understand new mechanisms that coordinate collective behaviors across great distances during tissue morphogenesis.

项目总结/摘要 神经管闭合缺陷和先天性心脏缺陷是常见的发育障碍， 每100名活产婴儿中分别有1名和1名死亡。在胚胎发育期间，细胞必须能够感知 指导和协调集体行为，以避免这些严重的畸形。虽然信号 在相邻细胞之间赋予方向性的途径是已知的，揭示了 数以千计的细胞已经停止了传递信息。幸运的是， 小型哺乳动物已经产生了因毛皮定向的区域特异性破坏而受到重视的动物。我 假设突变破坏了长距离的方向线索。本提案的目的是利用 自发突变，以确定哺乳动物表皮中的长距离方向线索。这项工作将脱落 了解复杂先天性疾病的病因，这将有助于子宫内治疗的发展 和人造器官的产生。 一个关键的分子途径，使细胞沟通方向的信息是平面细胞 极性（PCP）途径，没有它会出现严重的发育缺陷。核心路径包括三个 跨膜蛋白，形成位于细胞相对侧的不对称复合物。 通过细胞外正反馈和细胞内负反馈，这种不对称性得以传播 局部的，但整个组织的定向不对称的机制知之甚少。虽然核心 在果蝇中，通过正向遗传筛选鉴定途径组分，这种方法已经较少 成功地揭示了全球极性线索老鼠皮肤提供了一个理想的系统来识别远程 极性线索。膨胀的组织装饰着成千上万的毛囊（HF），这些毛囊都朝向 通过PCP驱动过程的相同方向。值得注意的是，我们发现背部皮肤是分开的 影响PCP信号方向的区域域。这些数据表明， 长距离提示协作以协调整个组织上的PCP和HF方向。为支持这一 想法，小型哺乳动物的自发突变产生的动物具有不同的，区域特异性的破坏， 毛囊方向为了研究这些突变是否会影响长程极性基因，我将鉴定 通过基因组测序确定致病突变，并表征突变对组织的影响 利用遗传学和图像分析来研究极性。然后，我将询问所鉴定的蛋白质作为 长距离极性线索在体内和器官型皮肤培养。在完成这项研究后，我们将确定 并了解在组织发育过程中协调远距离集体行为的新机制 形态发生