Interpreting Bone Morphogenetic Protein Gradients in Vertebrate Development

解释脊椎动物发育中的骨形态发生蛋白梯度

基本信息

批准号：
10677094
负责人：
Courtney Tello
金额：
$ 4万
依托单位：
STATE UNIVERSITY NEW YORK STONY BROOK
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：
2023-09-23 至 2025-09-22
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10677094
关键词：
Affect Animals Auxins Binding Blood Bone Morphogenetic Proteins Cell Fate Control Cell Transplantation Cells Characteristics Communication Complex Confocal Microscopy Critical Pathways Defect Development Disease Dose Elements Embryo Environment Exposure to Eye Abnormalities Fishes Fluorescent Probes Gastrointestinal Diseases Gene Activation Gene Expression Gene Expression Profiling Genes Genetic Genetic Transcription Germ Cells Germ Layers Goals Heart Abnormalities Heat-Shock Response Human Human Development Immune System Diseases Immune system In Situ Hybridization Kidney Knowledge Lead Length Light Link Location Malignant Neoplasms Mesoderm Mesoderm Cell Methods Microscopy Modeling Modification Molecular Outcome Output Paraxial Mesoderm Pathway interactions Pattern Positioning Attribute Pronephric structure Proteins Regenerative Medicine Reporter Research Resolution Role Signal Induction Signal Pathway Signal Transduction Signaling Protein Sjogren&apos s Syndrome Somites Specific qualifier value Syndrome System Systems Development Techniques Temporal bone structure Testing Time Tissues Transgenic Organisms Translating Transplantation Undifferentiated Vent Vertebrates Work Zebrafish body system cell behavior cell motility cell type combat gastrulation human disease in vivo insight intercellular communication migration morphogens mosaic mosaic analysis notochord novel organ growth overexpression progenitor short bone stem cells synergism transcription factor transgene expression zebrafish development

项目摘要

Project Summary/Abstract Animal development consists of complex cell coordination and rearrangement via intercellular communications. It is not well understood how progenitor cells interpret neighbor cell signals that impact cell fate decisions. One critical signaling element in animal development is morphogen signaling. Bone morphogenetic protein (BMP) acts as a morphogen to pattern the dorsoventral (DV) axis during vertebrate development, where BMP concentration gradients correlate with different cell identities. To study the mechanism in which progenitors interpret BMP gradients, I look to zebrafish development. Zebrafish are transparent during embryonic and larval stages, facilitating high resolution microscopy of cell fate change and migration patterns. Zebrafish share more than 70% of their genes with humans, making them excellent models for human development and disease research. Using zebrafish embryos, I will manipulate BMP concentration, progenitor cell location, and BMP signal duration within mesodermal progenitors and determine the morphogenetic outcome. In Aim 1 of this project, I will investigate the mechanism in which BMP signal coordinates its migration and cell fate gene expression that results in its characteristic morphogenetic output. With DV BMP morphogenetic signaling, cells may be positioned in embryonic domains where the microenvironment affects a binary fate switch. To answer my question, I will be using transgenic embryos to conditionally overexpress ectopic BMP signal. Transplanting transgenic cells into wildtype (WT) embryos prior to gastrulation stages will allow for the analysis of transgenic cell fate decisions within the WT environment. I will analyze time lapses of developing embryos containing these transgenic cells using spinning disk confocal microscopy. Cell fates will be genetically assessed using in situ hybridization and fate-specific fluorescent probes before, during and after gastrulation. In Aim 2, I will be determining the role that BMP signal duration has on cell fate decisions. I will be utilizing the Auxin Inducible Degron (AID) system to create specific durations of BMP signaling and BMP target gene expression in order to determine how signal duration impacts fate. Because a direct BMP target protein inhibits the transcription of cell fate specific transcription factors, I hypothesize that the duration of BMP signaling would change the amount of time that this target gene is present, allowing cell fate genes to bind when they normally would not be able to, thus altering cell fates. The combined use of cell autonomous reporter fish lines, transplantation, gene expression analysis, transgenic fish utilizing the AID system, and spinning disk confocal microscopy will allow me to test my hypotheses.

项目概要/摘要动物发育包括通过细胞间通讯进行的复杂细胞协调和重排。目前尚不清楚祖细胞如何解释影响细胞命运决定的邻近细胞信号。一动物发育中的关键信号元件是形态发生素信号传导。骨形态发生蛋白（BMP）在脊椎动物发育过程中，BMP 作为形态发生素来形成背腹 (DV) 轴，其中 BMP 浓度梯度与不同的细胞特性相关。研究祖细胞的作用机制解释 BMP 梯度时，我关注斑马鱼的发育。斑马鱼在胚胎和胚胎时期都是透明的幼虫阶段，促进细胞命运变化和迁移模式的高分辨率显微镜观察。斑马鱼分享它们70%以上的基因与人类相同，使它们成为人类发展的优秀模型疾病研究。使用斑马鱼胚胎，我将操纵 BMP 浓度、祖细胞位置，以及中胚层祖细胞内的 BMP 信号持续时间并确定形态发生结果。目标 1 在这个项目中，我将研究BMP信号协调其迁移和细胞命运基因的机制导致其特征形态发生输出的表达。通过 DV BMP 形态发生信号传导，细胞可能位于微环境影响二元命运转换的胚胎区域。来回答我的问题是，我将使用转基因胚胎来有条件地过度表达异位 BMP 信号。移植在原肠胚阶段之前将转基因细胞植入野生型（WT）胚胎中将允许分析转基因 WT 环境中的细胞命运决定。我将分析发育中的胚胎的时间流逝，其中包含这些转基因细胞使用转盘共聚焦显微镜。细胞命运将通过基因评估原肠胚形成之前、期间和之后的原位杂交和命运特异性荧光探针。在目标 2 中，我将确定 BMP 信号持续时间对细胞命运决定的作用。我将使用生长素诱导剂 Degron (AID) 系统创建特定持续时间的 BMP 信号传导和 BMP 靶基因表达，以便确定信号持续时间如何影响命运。因为直接 BMP 靶蛋白抑制 BMP 的转录细胞命运特异性转录因子，我假设 BMP 信号传导的持续时间会改变该目标基因存在的时间量，允许细胞命运基因在通常不会结合的情况下结合能够，从而改变细胞的命运。细胞自主报告鱼系的联合使用、移植、基因表达分析、利用 AID 系统的转基因鱼和转盘共聚焦显微镜将请允许我检验我的假设。