EXOGENOUS RECEPTOR PROBES OF SIGNALLING IN EMBRYOGENESIS

胚胎发生中信号传导的外源受体探针

• 批准号: 3329245
• 负责人: WILLIAM B BUSA
• 金额: $ 16.24万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1990
• 资助国家: 美国
• 起止时间: 1990-01-03 至 1993-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3329245
• 关键词: Xenopus; adenosine; adrenergic receptor; autoradiography; calcium; cell cell interaction; cell differentiation; cyclic AMP; cyclic GMP; diacylglycerols; early embryonic stage; embryo /fetus culture; embryogenesis; epidermal growth factor; fibroblast growth factor; in vitro fertilization; inositol phosphates; insulin receptor; mesoderm; messenger RNA; microinjections; muscarinic receptor; radioimmunoassay; receptor binding; receptor coupling; receptor expression; second messengers; serotonin receptor; vertebrate embryology

Both classical and modern studies point toward a central role for cell-cell signalling during early pattern formation events in the frog embryo, the preeminent experimental model of vertebrate development. Such signalling may involve secreted growth factor- like molecules and their plasma membrane receptors, a suggestion based on demonstrations that certain "classical" growth factors have close homologs in the maternal mRNA pool of the embryo and can induce isolated ectoderm to form mesodermal cell types. Our long-term interest is the role of signal transduction (receptor- mediated "second messenger" production) in the early signalling events which establish the body plan of the vertebrate embryo. This essentially biochemical step in the pattern formation process, lying as it does between two molecular genetic steps (on the one hand, the localization of determinants such as mRNAs in cytoplasm destined to compose "transmitting" cells, and on the other, the modification of gene expression which is the outcome of induction) has received little serious study, yet it is at precisely this chemical step where, we may suppose, many human teratogens (e.g., lithium, retinoic acid, etc.) are likely to exert their developmental effects. Unfortunately, familiar crude approaches for manipulating signalling systems -- microinjecting putative second messengers, treating with pharmacological inhibitors or activators of enzymes involved in signal transduction, and so forth -- have only rarely proven fruitful in studies of early development in intact embryos, where problems of long-term nonspecific toxicity, precise timing of stimulation (and its termination), and spatially localized delivery are routinely encountered. We have recently demonstrated the feasibility of a unique new approach to this issue: by injecting the frog zygote or early embryo with mRNA encoding an exogenous hormone receptor which is rapidly and functionally expressed in the blastula, we can install molecular "switches" in the embryo and use them to specifically stimulate or inhibit various second messenger systems (singly or in combinations) with a high degree of both spatial and temporal resolution. This novel approach provides us an unprecedented opportunity to understand the roles of these signalling events in early pattern formation, as demonstrated by our initial finding of profound effects on dorso-anterior differentiation in frog embryos expressing exogenous serotonin receptors linked to polyphosphoinositide hydrolysis and intracellular calcium mobilization.

古典研究和现代研究都指向 在早期模式形成事件中的细胞细胞信号传导 青蛙胚胎，脊椎动物的杰出实验模型 发展。 这种信号传导可能涉及分泌的生长因子 - 像分子及其质膜受体一样 基于某些“古典”增长因素的演示 在胚胎的母体mRNA库中具有密切的同源物， 可以诱导分离的外胚层形成中胚层细胞类型。 我们的 长期兴趣是信号转导的作用（受体 - 介导的“第二使者”制作）在早期信号中 建立脊椎动物胚胎的身体计划的事件。 在模式形成过程中，本质上是生化步骤， 躺在两个分子遗传步骤之间（在一个分子遗传步骤之间 手，诸如细胞质中mRNA之类的决定因素的定位 注定要组成“传输”细胞，另一方面是 基因表达的修饰，这是诱导的结果） 几乎没有认真研究，但正是 化学步骤，我们可以认为许多人类畸胎组（例如， 锂，视黄酸等）可能会施加 发展效果。 不幸的是，熟悉的操纵方法 信号系统 - 微注射假定的第二使者， 用酶的药理学抑制剂或活化剂处理 参与信号传输等等 - 很少有 在完整胚胎的早期发展研究中被证明是富有成果的 长期非特异性毒性的问题，精确的时机 刺激（及其终止），并在空间上定位 交货通常会遇到。 我们最近证明了 一种独特的新方法的可行性： 将青蛙合子或早期胚胎注射mRNA编码 外源激素受体，在功能上快速且功能 在胚胎中表达，我们可以安装分子 胚胎中的“开关”，并使用它们专门刺激或 抑制各种第二会系统（单一或 组合）具有高度的空间和时间。 解决。 这种新颖的方法为我们提供了前所未有的 了解这些信号事件在中的作用的机会 早期模式形成，如我们最初的发现 对青蛙胚胎中多索 - 验分化的深刻影响 表达与 多磷酸肌醇水解和细胞内钙 动员。