TISSUE SPECIFIC CONTROL OF CELL PROLIFERATION

细胞增殖的组织特异性控制

• 批准号: 6173777
• 负责人: WAYNE N. FRANKEL
• 金额: $ 31.74万
• 依托单位: JACKSON LABORATORY
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-06-01 至 2002-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6173777
• 关键词: biomarker; cell growth regulation; cell proliferation; cell type; chromosome walking; developmental genetics; gene expression; gene mutation; gene targeting; genetic mapping; genetically modified animals; laboratory mouse; neural plate /tube; nucleic acid sequence; phenotype; proliferating cell nuclear antigen; vertebrate embryology

DESCRIPTION (Adapted from investigator's abstract): An essential component of tissue morphogenesis, function, and homeostasis is the differential control of cell proliferation in the different cell types that comprise a tissue. The long term goal of the proposed studies is to understand the molecular mechanisms underlying such cell type-specific controls of cell proliferation. As a starting point toward the identification of genes that participate in the cell type-specific regulation of cell proliferation, Gossler has chosen to focus on mouse mutations that affect the proliferation of particular cell types or tissues. In this proposal, the mouse mutation under study is Pintail (Pt). Pintail was isolated from the offspring of mice treated with the carcinogen methylcholantrene. It is a semidominant mutation on chromosome 4 and it specifically affects the proliferation of notochord cells in a dose dependent manner. A descriptive study of Pt, published nearly 40 years ago, showed that the frequency of mitotic cells in the notochord was reduced to approximately 50% and 30%, of normal values in, respectively, heterozygotes and homozygotes. As a result, the notochord is significantly smaller or completely absent in the posterior portion of the embryo; this in turns generates skeletal abnormalities. The proposed experiments are divided into four specific aims. Specific Aim 1 further explores the Pt phenotype. First, an analysis of BrdU incorporation into notochord cells of mutant and wild type embryos will be performed to define the G1, S, G2, and M parameters of the cell cycle in Pt mutant cells. In addition, an expression analysis will be conducted on markers for defined phases of the cell cycle. Second, chimeras will be constructed by the injection of Pt ES cells into wild type blastocysts to assess the cell autonomy of the Pt mutation. Specific Aim 2 proposes to establish a fine scale genetic map of the Pintail mutation using at least 2000 backcross animals generated from two different intersubspecific backcrosses. Using the genetic map and chromosomal walking techniques, a physical map and a BAC contig will be constructed for the Pt critical interval. The Pt gene will be identified in Specific Aim 3 using the candidate gene approach, cDNA selection, and/or sequencing of a <50 kb critical region. The authenticity of the Pt gene will be verified by expression analyses, sequencing, and appropriate transgenic or knockout experiments. In Specific Aim 4, transgenic approaches will be undertaken to functionally analyze the pintail gene and its role in cell type specific control of cell proliferation.

描述（改编自研究者摘要）：一种基本成分 组织形态发生、功能和体内平衡的差异是 控制不同细胞类型中的细胞增殖， 组织. 拟议研究的长期目标是了解 这种细胞类型特异性控制细胞的分子机制 增殖 作为识别基因的起点， 参与细胞增殖的细胞类型特异性调节， Gossler选择关注影响增殖的小鼠突变， 特定的细胞类型或组织。 在这个提议中， Pintail（Pt）。 针尾鱼是从 用致癌物质甲基胆甾烯治疗的小鼠。 这是一个半显性的 4号染色体上的突变，它特异性地影响 脊索细胞以剂量依赖的方式。 Pt的描述性研究， 近40年前发表的一项研究表明， 脊索减少到正常值的约50%和30%， 分别为杂合子和纯合子。 因此，脊索是 明显更小或完全没有在后面的部分 胚胎;这反过来又产生骨骼异常。 拟议的实验分为四个具体目标。 具体目标 1进一步探讨了Pt表型。 首先，对BrdU进行分析 将突变体和野生型胚胎掺入脊索细胞中， 用于定义Pt中细胞周期的G1、S、G2和M参数 突变细胞 此外，还将对以下内容进行表达分析： 用于细胞周期的限定阶段的标记物。 第二，嵌合体将 通过将Pt ES细胞注射到野生型胚泡中构建， 评估Pt突变的细胞自主性。 具体目标2建议， 至少使用以下方法建立针尾突变的精细遗传图谱： 从两个不同亚种间产生的2000只回交动物 回交 利用遗传图谱和染色体步移技术， 物理图谱和BAC重叠群将被构建用于Pt关键 interval. Pt基因将在特定目标3中使用 候选基因方法、cDNA选择和/或测序<50 kb的 关键区域。 Pt基因的真实性将通过 表达分析、测序和适当的转基因或敲除 实验 在具体目标4中，将采用转基因方法， 分析pintail基因的功能及其在细胞类型特异性 控制细胞增殖。