The Role of Calcium Transients in Fertilization and Early Embryogenesis in C. elegans

钙瞬变在秀丽隐杆线虫受精和早期胚胎发生中的作用

基本信息

批准号：
10654250
负责人：
Karen S Kim Guisbert
金额：
$ 43.04万
依托单位：
FLORIDA INSTITUTE OF TECHNOLOGY
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-06-01 至 2026-05-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10654250
关键词：
Academic Research Enhancement Awards Address Animals Biological Models Caenorhabditis elegans Calcium Calcium Oscillations Calcium Signaling Calcium Spikes Cell Communication Cell division Cells Communication Competence Complement Complex Consumption Cytology Data Defect Dependence Development Developmental Biology Dyes Education Educational process of instructing Embryo Embryo Research Embryonic Development Event Fertility Fertilization Florida Genetic Germ Cells Goals Gonadal structure Human Human Development Image In Vitro Infertility Injections Knowledge Learning Marriage Meiosis Microscopy Mitotic Molecular Nature Neurons Oocytes Optics Organism Ovulation Pattern Phase Phenotype Physiology Process RNA interference screen Regulation Reporter Reporting Reproductive Medicine Research Research Infrastructure Research Project Grants Role Series Signal Pathway Signal Transduction Slice Specificity Spontaneous abortion Students System Techniques Technology Testing Time Visualization biochemical tools calcium indicator cell type course redesign critical period design egg experimental study gastrulation improved in vivo innovation insight knock-down laboratory experience model organism novel prevent response sperm cell student participation tool undergraduate research experience undergraduate student

项目摘要

Project Summary Human development begins with an internal fertilization event. However, internal fertilization is notoriously difficult to visualize in vivo. Taking advantage of the universally conserved calcium response that is a hallmark of sperm-egg fusion in all species, we have adapted the genetically encoded calcium indicator jGCaMP7s to fluorescently indicate the moment of fertilization inside a living animal, C. elegans. Our system produces a robust signal that recapitulates the previously reported, biphasic nature of the calcium wave and has no deleterious effects on worm physiology or fecundity. This reporter is a powerful new tool to facilitate our long-term goal to define cell-cell communication and coordination during the events of the oocyte-to-embryo transition and early embryogenesis. The rationale is that visualization of the calcium transients, in conjunction with genetic and molecular tools, will allow an unprecedented opportunity to dissect early embryogenesis inside a living animal. Completion of Specific Aim 1 will define novel intercellular regulation and coordination that occurs during the oocyte-to- embryo transition by examining our reporter in a background that forces both a fertilization-competent and a fertilization-incompetent oocyte to be ovulated at the same time. Specific Aim 2 will establish the cell-type specificity and timing of calcium transients during early embryogenesis that have never before been observed in C. elegans. Specific Aim 3 will incorporate a targeted RNAi screen for OET regulators into a redesigned course-based undergraduate research experience (CURE) lab. This will synergistically propel OET research and improve undergraduate Developmental Biology education at Florida Tech. The central innovation of this proposal is the marriage of superior calcium imaging during early embryogenesis combined with a model system that has superlative genetics and a well-ordered gonad. This proposed research is significant because it will enable new insights into the timing and regulation of the OET and early embryogenesis in an intact living animal and could led to advances in reproductive medicine. Importantly, this proposal will satisfy key objectives of the R15 mechanism and the Notice of Special Intent (NOT-HD-19-036) by 1) significantly enhancing exposure of students to developmental biology research and 2) strengthening the research infrastructure at the Florida Institute of Technology.

项目概要人类发育始于体内受精事件。然而，内部众所周知，受精在体内很难可视化。充分发挥普遍优势保守的钙反应是所有物种精卵融合的标志，我们使基因编码的钙指示剂 jGCaMP7s 适应荧光表示活体动物秀丽隐杆线虫体内受精的时刻。我们的系统产生一个强大的信号，概括了先前报道的双相性质钙波，对蠕虫生理或繁殖力没有有害影响。这记者是一个强大的新工具，可以促进我们定义细胞的长期目标卵母细胞向胚胎转变过程中的沟通和协调早期胚胎发生。基本原理是钙瞬变的可视化，与遗传和分子工具相结合，将提供前所未有的机会解剖活体动物体内的早期胚胎发生。完成具体目标 1 将定义发生在卵母细胞到-的过程中的新型细胞间调节和协调通过在迫使双方有受精能力和无受精能力的卵母细胞同时排卵时间。具体目标 2 将确定细胞类型特异性和钙瞬变时间在早期胚胎发生过程中，这种现象以前从未在秀丽隐杆线虫中观察到过。具体目标 3 将针对 OET 调节剂的靶向 RNAi 筛选纳入重新设计了基于课程的本科生研究体验（CURE）实验室。这将协同推动 OET 研究并提高本科生发育生物学佛罗里达理工学院的教育。该提案的核心创新是将早期胚胎发生期间的卓越钙成像与模型系统相结合拥有最高级的遗传学和秩序良好的性腺。这项拟议的研究意义重大因为它将让我们对 OET 的时间安排和监管以及早期完整活体动物的胚胎发生可能导致生殖技术的进步药品。重要的是，该提案将满足 R15 机制的关键目标特别意图通知 (NOT-HD-19-036) 1) 显着提高学生进行发育生物学研究和2）加强研究佛罗里达理工学院的基础设施。