Dynamic polarity mechanisms controlling stem cell asymmetry during tissue development

组织发育过程中控制干细胞不对称的动态极性机制

• 批准号: 9756654
• 负责人: Andrew D Muroyama
• 金额: $ 6.12万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-08-01 至 2021-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9756654
• 关键词: Acute; Animal Model; Arabidopsis; Automobile Driving; Biological Assay; Cell Polarity; Cell Size; Cells; Complex; Confocal Microscopy; Coupled; Data; Daughter; Defect; Development; Epidermis; Equilibrium; Feedback; Future; Generations; Genetic; Growth; Health; Heterogeneity; Homeostasis; Human; Investigation; Link; Logic; MAP Kinase Gene; Malignant Neoplasms; Mediating; Microscopy; Microtubules; Modeling; Molecular; Molecular Genetics; Mouse-ear Cress; Multipotent Stem Cells; Optics; Organogenesis; Pathway interactions; Pattern; Phenotype; Plant Leaves; Plants; Proteins; Proteomics; Reporter; Research; Resolution; Stem cells; System; Testing; Time; Tissues; Transgenic Organisms; Work; base; cell behavior; cell growth; cell type; daughter cell; design; developmental disease; experimental study; high resolution imaging; human disease; in vivo; in vivo imaging; mutant; novel; self-renewal; stem cell division; synthetic biology; time use; tool; transcriptome

PROJECT SUMMARY/ABSTRACT Cell polarity is necessary to pattern tissues and is essential in stem cells to generate cellular diversity during development and homeostasis. Accordingly, polarity defects can drive developmental disorders and diseases, including cancer. Previous studies have elucidated molecular mechanisms that generate polarity and break symmetry in single cells. How these mechanisms are integrated to generate complex tissues remains poorly understood. The stomatal lineage in the Arabidopsis leaf epidermis is an ideal system to quantitatively determine how intrinsic cell polarity is transmitted to daughter cell asymmetry due to its well-characterized developmental trajectory, optical accessibility, and genetic tractability. In this proposal, I will utilize the stomatal lineage to 1) test how a novel microtubule-dependent inhibitory loop utilizes conserved logic to generate polarity in stem cells and 2) delineate polarity-dependent mechanisms that drive daughter cell asymmetry. To accomplish the proposed aims, I have developed novel in vivo tools to manipulate polarity in the stomatal lineage with high temporal and subcellular precision. In Aim 1, I will use targeted depletion, molecular genetics, and high-resolution imaging to test how mutual inhibition between microtubules and cortical polarity proteins generate a single polarity axis in multipotent stem cells. Additionally, I will use quantitative in vivo imaging to experimentally test how integrating an inhibitory circuit with a parallel MAPK-dependent positive feedback loop increases polarity robustness. In Aim 2, I will leverage the strengths of the stomatal lineage to determine how stem cell polarity regulates differential daughter cell size. By acutely manipulating polarity and tracking resulting daughter cell size, growth, and fate decisions using long-term time-lapse microscopy in vivo, I will directly test how polarity regulates downstream asymmetry. Furthermore, by identifying novel polarity regulators in the stomatal lineage, I will identify molecular pathways that link polarity to differential daughter growth. Completion of the proposed aims will test polarity models during multicellular development and will identify the mechanisms linking polarity to stem cell asymmetry. Elucidation of polarity mechanisms in an evolutionarily divergent species has broad implications for polarity models and may define synthetic strategies to correct polarity defects in human disease states.

项目摘要/摘要 细胞极性是组织构型所必需的，也是干细胞产生细胞多样性所必需的。 在发育和动态平衡期间。因此，极性缺陷会导致发育障碍和 疾病，包括癌症。以前的研究已经阐明了产生极性和 打破单个单元格的对称性。这些机制如何整合以产生复杂的组织仍然存在 人们对此知之甚少。拟南芥叶表皮气孔谱系是定量测定的理想系统 确定固有的细胞极性如何传递给子细胞的不对称性，这是由于其特征良好 发育轨迹、视觉可及性和遗传可控性。在这个提案中，我将利用气孔 1)测试新的微管依赖抑制环如何利用守恒逻辑产生极性 和2)描绘了驱动子代细胞不对称的极性依赖机制。要完成 提出的目标，我已经开发了新的活体工具来操纵气孔谱系中的极性 时间和亚细胞精度。在目标1中，我将使用定向耗竭、分子遗传学和高分辨率 成像以测试微管和皮质极性蛋白之间的相互抑制如何产生单个 多能干细胞中的极轴。此外，我将使用体内定量成像来进行实验测试 如何将抑制电路与依赖MAPK的并行正反馈环路集成在一起增加极性 健壮性。在目标2中，我将利用气孔谱系的优势来确定干细胞的极性 调节不同的子细胞大小。通过敏锐地操纵极性和跟踪所得的子细胞大小， 在活体中使用长期延时显微镜，我将直接测试极性是如何调节的 下游不对称。此外，通过在气孔谱系中识别新的极性调节器，我将 确定将极性与不同的子代生长联系起来的分子途径。完成拟议的目标 将在多细胞发育过程中测试极性模型，并将确定将极性与茎 细胞不对称。阐明进化上不同物种的极性机制具有广泛的意义 用于极性模型，并可定义纠正人类疾病状态中的极性缺陷的合成策略。