Intracellular pH Dynamics in Zebrafish Cranial Neural Crest Development

斑马鱼颅神经嵴发育的细胞内 pH 动态

• 批准号: 10704106
• 负责人: Cambria Chou-Freed
• 金额: $ 4.51万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-09-05 至 2025-09-04
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10704106
• 关键词: Address; Behavior; Biology; Cell Adhesion; Cell Fate Control; Cell membrane; Cells; Cephalic; Chick; Chondrocytes; Craniofacial Abnormalities; Data; Defect; Development; Disease; Drosophila genus; Embryo; Embryonic Development; Exhibits; Focal Adhesion Kinase 1; Future; Genetic; Genetic Transcription; Health; Human; Image; Intestines; Ions; Modeling; Molecular; Mus; Neural Crest; Neural Crest Cell; Odontoblasts; Organism; Osteocytes; Ovarian Follicle; Paraxial Mesoderm; Population; Proteins; Regulation; Reporter; Reporting; Role; Specific qualifier value; System; Talin; Testing; Time; Tissues; Zebrafish; blastomere structure; cell behavior; cell fate specification; cell motility; cell type; cofilin; craniofacial; craniofacial development; embryo cell; embryonic stem cell; epithelial to mesenchymal transition; forging; in vivo; innovation; insight; interest; lens; live cell imaging; migration; non-genetic; novel; pharmacologic; protonation; ratiometric; sensor; single-cell RNA sequencing; stem cells; tissue repair; vertebrate embryos

PROJECT SUMMARY/ABSTRACT Intracellular pH (pHi) dynamics are a critical regulator of cell fate changes. Our lab and others demonstrated that an increase in pHi is necessary for the differentiation and lineage specification of multiple cell types, including mouse embryonic stem cells, Drosophila ovarian follicle stem cells, mouse intestinal stem cells, and chick paraxial mesoderm. Our lab has also shown how pHi dynamics regulate cell adhesion and migration behaviors by modifying the protonation state of pH-sensing proteins, including -catenin, cofilin, talin, and focal adhesion kinase. However, the mechanisms underlying pHi regulation of cell fate changes remain unresolved. Furthermore, our understanding of pHi dynamics during embryonic development in vivo is limited, largely due to lack of appropriate models. To address this, I generated a novel system for interrogating the functional significance of pHi dynamics during embryonic development in vivo using zebrafish, a genetically tractable organism suited for in vivo live cell imaging of embryos. Using this new model, I will determine the role of pHi dynamics in the development of cranial neural crest (NC), a highly conserved vertebrate embryonic cell population that gives rise to diverse cell types, including chondrocytes, osteocytes, and odontoblasts. Many cell behaviors involved in cranial NC development are regulated by pHi dynamics in other cell types, including lineage specification, cell migration, and epithelial to mesenchymal transition. Thus, cranial NC represents an ideal model for addressing the gaps in our understanding of pHi dynamics during embryonic development in vivo. By following early NC cells through the differentiation of cranial lineages, I will test the central hypothesis that pHi dynamics regulate cranial NC development, at the stage of delamination, migration, or lineage specification. In Aim 1, I will resolve spatial and temporal pHi dynamics during zebrafish cranial NC development by in vivo live cell imaging. My preliminary data indicate a higher pHi in migratory compared with premigratory cranial NC cells. In Aim 2, I will experimentally perturb pHi in zebrafish NC cells through pharmacologic and genetic modulation of plasma membrane ion transporters and determine the effect on cranial NC cell behaviors and transcription, thus establishing the functional significance of pHi dynamics during zebrafish cranial NC development. My findings have promise to reveal new insight on the cellular and molecular factors controlling craniofacial development, with important implications for human congenital diseases and tissue repair.

项目摘要/摘要 细胞内pH（PHI）动力学是细胞命运变化的关键调节剂。 对于多种细胞类型的差异率和谱系规范，PHI的增加是必需的 包含小鼠胚胎干细胞，果蝇卵巢卵泡干细胞，小鼠肠干细胞，以及 雏鸡中胚层。我们的实验室还显示了PHI动力学 通过修改pH传感蛋白的质子化状态，包括-catenin，cofilin，talin和焦点 但是，粘附激酶。 此外，我们对体内胚胎开发过程中PHI动态的理解是有限的，在很大程度上是由于 缺乏适当的模型来解决这个问题 斑马鱼在体内胚胎发育过程中Phi动力学的意义，斑马鱼是一种遗传学的 适合胚胎体内活细胞成像的生物体。 颅神经rest（NC）的发展，一种高度保守的脊椎动物胚胎细胞 产生各种细胞类型的种群，包括骨细胞，骨细胞和牙胶细胞 颅NC发育中涉及的细胞行为是通过其他细胞类型的PHI动力学定制的，包括 谱系规范，细胞迁移和上皮向间充质转变。 解决我们对胚胎发展过程中PHI动力学的理解差距的理想模型 体内通过颅骨的差异来遵循早期的NC细胞，我将测试中心 假设PHI动力学常规颅NC在分层阶段， 在AIM 1中迁移或谱系规范。 斑马鱼NC通过体内活细胞成像开发。 与AIM 2中的颅内NC细胞相比 NC细胞通过质膜离子转运蛋白的药理和遗传调节，并确定 对颅NC细胞行为和转录的影响，从而确立了PHI的功能意义 斑马鱼颅NC开发期间的动态。 控制颅面发育的细胞和分子因素，对人类的重要性很重要 先天性疾病和组织修复。