Cytokeratin Organization and Function in Early Development

早期发育中的细胞角蛋白组织和功能

• 批准号: 8905022
• 负责人: Michael Klymkowsky
• 金额: $ 24.34万
• 依托单位: University of Colorado at Boulder
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 1989
• 资助国家: 美国
• 起止时间: 1989-08-01 至 1994-01-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=8905022&HistoricalAwards=false
• 关键词: Cytokeratin; Organization; Function; Early; Development

Cytokeratin filaments from a major cytoskeletal system in the Xenopus oocyte and early embryo. Using wholemount immunocytochemistry Dr. klymkowsky has discovered a dramatic asymmetry in the organization of cytokeratin filaments in both the oocyte and early embryo. During oocyte maturation cytokeratin filaments break down. Cytokeratin breakdown appears to be regulated post-translationally and is induced by maturation promoting factor (MPF). Dr. Klymkowsky will determine whether MPF acts directly or indirectly to disrupt cytokeratin organization using an in vitro assay system. With fertilization cytokeratin filaments reform; this asymmetrically organized embryonic cytokeratin system persists until the end of gastrulation. To define the function of cytokeratin filaments in the embryo, he will develop new monoclonal anticytokeratin antibodies. The epitopes of these antibodies will be mapped on the cytokeratin protein and its effects on cytokeratin filament integrity and embryonic development will be determined. This will enable him to define regions of the cytokeratin molecule involved in cytokeratin filament organization and function. Dr. Klymkowsky has already identified two monoclonal anticytokeratin antibodies that specifically disrupt normal gastrulation in Xenopus. To define further the mechanism by which these antibodies act, he will use light and electron microscopy to characterize the movement and morphology of cells during gastrulation and immunocytochemistry to examine cytokeratin organization within the embryo. Injection of antibodies into specific blastomeres will be used to determine the regions of the gastrulating embryo affected by these antibodies. Together these studies offer an opportunity for defining the functions of cytokeratins, understanding how their organization if controlled and for furthering our understanding of the major morphogenetic event of vertebrate development, gastrulation. %%% One of the most dramatic events during the maturation of the egg in the amphibian, Xenopus is the reorganization of the subcellular cytokeratin-type intermediate filaments. The resulting mature egg is asymmetric and this asymmetry defines the axis of the mature organism. Clearly the events leading to the setting up of this polarity are key events in development.

爪蟾卵母细胞和早期胚胎中主要细胞骨架系统的细胞角蛋白丝。利用全量免疫细胞化学，klymkowsky博士发现了卵母细胞和早期胚胎中细胞角蛋白丝组织的显著不对称性。在卵母细胞成熟过程中，细胞角蛋白丝分解。细胞角蛋白的分解似乎在翻译后受到调控，并由成熟促进因子（MPF）诱导。Klymkowsky博士将使用体外分析系统确定MPF是否直接或间接地破坏细胞角蛋白组织。受精后细胞角蛋白细丝重组；这种不对称组织的胚胎细胞角蛋白系统一直持续到原肠胚形成结束。为了确定细胞角蛋白丝在胚胎中的功能，他将开发新的单克隆抗细胞角蛋白抗体。这些抗体的表位将被定位在细胞角蛋白上，并将确定其对细胞角蛋白丝完整性和胚胎发育的影响。这将使他能够确定参与细胞角蛋白丝组织和功能的细胞角蛋白分子区域。克莱姆科夫斯基博士已经发现了两种单克隆抗细胞角蛋白抗体，它们能特异性地破坏非洲爪蟾正常的原肠胚形成。为了进一步确定这些抗体的作用机制，他将使用光学和电子显微镜来描述原肠胚形成过程中细胞的运动和形态，并使用免疫细胞化学来检查胚胎内的细胞角蛋白组织。将抗体注射到特定的卵裂球中，将用于确定受这些抗体影响的原肠胚的区域。总之，这些研究提供了一个机会来定义细胞角蛋白的功能，了解它们的组织是如何被控制的，并进一步加深我们对脊椎动物发育的主要形态发生事件——原肠胚形成的理解。两栖动物爪蟾卵子成熟过程中最引人注目的事件之一是亚细胞角蛋白型中间丝的重组。由此产生的成熟卵子是不对称的，这种不对称定义了成熟生物体的轴线。很明显，导致这种极性形成的事件是发展中的关键事件。