Cell Cycle Regulation of Cell Fate and Morphogenesis in D. rerio

斑马鱼细胞命运和形态发生的细胞周期调控

基本信息

批准号：
10627845
负责人：
Samantha Stettnisch
金额：
$ 3.81万
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：
2022
资助国家：
美国
起止时间：
2022-05-19 至 2025-05-18
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10627845
关键词：
Adopted Affect Anterior Behavior Biosensor Brachyury protein CDK2 gene CDK4 gene CDKN1C gene Cell Cycle Cell Cycle Arrest Cell Cycle Inhibition Cell Cycle Regulation Cell Fate Control Cell Transplantation Cells Complex Confocal Microscopy Cues Cyclin-Dependent Kinase Inhibitor Cyclin-Dependent Kinases Data Development Disease Disseminated Malignant Neoplasm Embryo Exclusion Floor G1 Arrest Gene Expression Genes Goals Invaded Knowledge Malignant Neoplasms Mediating Modeling Morphogenesis Movement Organism Outcome Phase Play Population Process Proliferating Publishing Regulation Research Role Signal Pathway Signal Transduction Specific qualifier value Structure Testing Tissues Transcriptional Regulation Transgenic Organisms Up-Regulation Work Zebrafish cancer cell cell behavior cell fate specification cell motility cellular imaging convergent extension developmental disease gene regulatory network in vivo in vivo Model insight intercalation knock-down migration mutant new therapeutic target notch protein notochord notochord development novel novel therapeutics overexpression prevent progenitor single-cell RNA sequencing stem cells transcription factor zebrafish development

项目摘要

PROJECT SUMMARY / ABSTRACT Complex morphogenetic processes are required for proper organismal development. These morphogenetic processes require various combinations of cell migration, proliferation, invasion, and fate acquisition. The coordination of these behaviors must be tightly regulated, as dysregulation of these processes can lead to developmental disorders and disease states such as cancer. To study the regulation of complex morphogenetic process we turn to zebrafish development. In zebrafish, morphogenesis of midline tissue structures such as the notochord, floor plate, and hypochord drive axis elongation of the developing embryo. These tissues are derived from a population of progenitors residing in the tailbud known as midline progenitor cells (MPCs). MPCs undergo a morphogenetic process called convergent extension (CE) to give rise to the notochord. During CE, adjacent MPCs migrate and intercalate between one another to form the notochord. The decision of MPCs to adopt a notochord, floor plate, or hypochord fate is based on local signaling cues such as Wnt and Notch. While these signaling pathways have been shown to regulate morphogenetic cell behaviors, there is growing evidence to suggest that the cell cycle can also modulate cell behaviors. However, the mechanisms by which cell cycle state dictates cell behavior and cell fate during tailbud morphogenesis remain unclear. I will address this gap in knowledge and elucidate the relationship between cell cycle state and cell fate/morphogenesis during development using CE of the zebrafish notochord during tailbud morphogenesis as a model. Published data from my lab and others show notochord progenitor cells are in G1, while floor plate and hypochord progenitors can be found in all phases of the cell cycle. Furthermore, these notochord progenitors undergo CE in G1 and reenter the cell cycle after joining the notochord, suggesting that G1 arrest facilitates this morphogenetic process and notochord fate acquisition. In Aim 1 of this project, I will perturb G1 arrest specifically in the MPCs and determine the effects on CE. Using transgenic zebrafish lines, including a CDK activity biosensor to establish cell cycle state, and spinning disk confocal microscopy, I will time-lapse image these cell cycle perturbed embryos and quantify CE and fate acquisition. In Aim 2, I will explore the gene regulatory network (GRN) responsible for inducing G1 arrest in the MPCs, with a focus on the T-box transcription factor Brachyury (tbxta). Published data show tbxta to be indispensable to notochord formation and moreover, preliminary data from our lab show knockdown of tbxta drives notochord progenitors to cycle and be excluded from the notochord, adopting a floor plate or hypochord fate. The combination of a CDK activity biosensor, cell cycle perturbation constructs, midline directed cell transplantation, and spinning disk confocal microscopy will allow me to test my hypotheses thoroughly and rigorously.

项目摘要 /摘要适当的生物发育需要复杂的形态发生过程。这些形态发生过程需要各种细胞迁移，增殖，入侵和命运的组合。这这些行为的协调必须严格调节，因为这些过程的失调可能导致发育障碍和疾病状态，例如癌症。研究复杂的调节形态发生过程我们转向斑马鱼的发展。在斑马鱼中，中线组织的形态发生诸如发育中胚胎的脊索，地板板和次音驱动轴伸长等结构。这些组织源自位于被称为中线祖细胞的尾巴中的祖细胞种群细胞（MPC）。 MPC经历了称为收敛扩展（CE）的形态发生过程，以产生脊索。在CE期间，相邻的MPC彼此之间迁移和插入以形成Notochord。 MPC采用脊索，地板板或次体命运的决定是基于局部信号提示例如Wnt和Notch。尽管这些信号通路已显示用于调节形态发生细胞行为，越来越多的证据表明细胞周期也可以调节细胞行为。然而，细胞周期状态在尾腹形态发生过程中决定细胞行为和细胞命运的机制保持不清楚。我将在知识方面解决这一差距，并阐明细胞周期状态与在发育过程中使用斑马鱼脊索CE发育过程中的细胞命运/形态发生。形态发生为模型。来自我的实验室的发布数据和其他数据显示，固定核祖细胞在G1中，虽然在细胞周期的所有阶段都可以找到地板板和次体祖细胞。此外，这些 Notochord祖细胞在G1中经历CE，并在加入Notochord后重新进入细胞周期，这表明 G1停滞促进了这种形态发生过程和努力命运的获取。在该项目的AIM 1中，我将扰动G1在MPC中专门逮捕，并确定对CE的影响。使用转基因斑马鱼线，包括CDK活性生物传感器以建立细胞周期状态和旋转磁盘共聚焦显微镜，我将延时图像这些细胞周期扰动胚胎并量化CE和命运的采集。在AIM 2中，我会探索负责在MPC中诱导G1停滞的基因调节网络（GRN），重点是 T-box转录因子Brachyury（TBXTA）。已发布的数据显示，tbxta对于notochord是必不可少的此外，我们实验室的初步数据显示了tbxta的敲低驱动器固定祖细胞循环并从脊索中排除，采用地板板或次体命运。结合 CDK活性生物传感器，细胞周期扰动构建体，中线定向细胞移植和旋转磁盘共聚焦显微镜将使我能够彻底，严格地测试我的假设。