EXPRESSION OF ENGRAILED IN IDENTIFIED NEURONS OF CNS

目前的工作是确定以下方面作用的总体努力的一部分 同源框基因在神经元特异性发育和维持中的作用 在中枢神经系统中。与健康相关的工作方面有 在越来越多的研究中容易得到认可，这些研究表明 在大脑和脊髓以及一些非神经元组织中存在缺陷， 当同源盒基因在发育过程中被破坏时。尽管 同源盒中断导致的中枢神经系统缺陷通常在 相当广泛地说，缺陷最终必须驻留在 单个神经元的表型，以及更进一步，由改变的 特定结构基因的表达。同源盒基因通常是 被认为是发育基因。尽管如此，一些人还是继续表达 进入成年期，因此推测它们在 维持神经元结构、连通性或 功能。第一个具体目标是描述 血吸虫成虫中枢神经系统的聚集表达。积极向上的是 基因的蛋白质产物，并将使用 识别蛋白质同源结构域的单抗(单抗4D9)。 将检查中枢神经系统的免疫标记神经节以评估刻板印象 标签的位置、范围和相对于地标的位置。这个 第二个特定目标是检验累加表达式是 特定类型，表达发生在神经元间，而不是传出 线性相关的神经元群。我们将检查分段同源 中线神经元群，每个神经元组成已识别的 胚胎干细胞。对这一假设的直接检验将由 双标记实验：单抗4D9将与逆行联合使用 标记或细胞内注射染料到已识别的神经元中 中线组。我们将评估不同标签之间的重叠 神经元类型。类型特异性的确凿证据将是 通过对累加正负的定量分析得到 血统组内的神经元。第三个具体目标是 描述胚胎发育过程中渐进式表达的特征。我们 假设渐变表达式在一个可定义的空间和 胚胎发育过程中的时间模式，以产生成熟模式。二 提出了多个层次的调查，一个是在人口层面：我们将建立 通过计数免疫标记和未标记的细胞 中线组以5%为增量，胚胎发育率从30%提高到100%。 我们的第二个层次的研究将以细胞表达为特征 决议。我们将追踪干细胞和 包含最新出生的后代的独特的解剖图形。 这些数据加上对细胞周期时间的估计，将使 表达的开始和偏移相对于时间的特征 出生。

The present work is part of a general effort to determine the role of homeobox genes in the development and maintenance of neuronal specificity in the central nervous system. Health related aspects of the work are readily recognized in the growing number of studies that show specific defects in brain and spinal cord, and in a number of non-neuronal tissues, when homeobox genes have been disrupted during development. Although the CNS defects resulting from homeobox disruption are typically described in rather broadly, the defects must ultimately reside in alterations in the phenotype of individual neurons, and further, result from the altered expression of particular structural genes. Homeobox genes are commonly regarded as developmental genes. Nonetheless, some continue expression into adulthood, and it is therefore speculated that they have a role in maintaining the integrity of neuronal structure, connectivity, or function. The first specific aim is to characterize the pattern of Engrailed expression in the adult CNS of Schistocerca. Engrailed is the protein product of the engrailed gene and will be detected using a monoclonal antibody (MAb 4D9) that recognizes the protein's homeodomain. Immunolabeled ganglia of the CNS will be examined to assess the stereotypy of the labeling, its extent and its location relative to landmarks. The second specific aim is to test the hypothesis that Engrailed expression is type-specific, with expression occurring interneurons but not efferents of lineally related neuronal groups. We will examine segmentally homologous midline groups of neurons which each comprise the progeny of an identified embryonic stem cell. A direct test of the hypothesis will be made by double labeling experiments: MAb 4D9 will be used together with retrograde labeling or intracellular injection of dye into identified neurons in the midline groups. We will assess the overlap between labels in different neuronal types. Corroborating evidence for type-speCificity will be obtained by a quantitative analysis of Engrailed-positive and -negative neurons within the lineage groups. The third specific aim is to characterize the Engrailed expression during embryonic development. We hypothesize that Engrailed expression evolves in a definable spatial and temporal pattern during embryogenesis to yield the mature pattern. Two levels of inquiry are proposed, one at population level: We will construct a population profile by counting immunolabeled and unlabeled cells in the midline groups at 5% increments from 30% to 100% of embryonic development. Our second level of inquiry will characterize expression with cellular resolution. We will trace expression within the stem cell and in a distinct anatomical figure that contains the most recently born progeny. These data together with estimates of cell cycle time will allow a characterization of the onset and offset of expression relative to time of birth.