Biology of a New DAP12 Associated Receptor Family

新 DAP12 相关受体家族的生物学

• 批准号: 6330740
• 负责人: William E Seaman
• 金额: $ 27.83万
• 依托单位: NORTHERN CALIFORNIA INSTITUTE/RES/EDU
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-05-01 至 2006-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6330740
• 关键词: antireceptor antibody; cell migration; complementary DNA; cytokine; gene targeting; genetically modified animals; laboratory mouse; ligands; macrophage; monoclonal antibody; nitric oxide; polymerase chain reaction; protein structure function; receptor; receptor binding; receptor expression; antireceptor antibody; cell migration; complementary DNA; cytokine; gene targeting; genetically modified animals; laboratory mouse; ligands; macrophage; monoclonal antibody; nitric oxide; polymerase chain reaction; protein structure function; receptor; receptor binding; receptor expression

DESCRIPTION (provided by the applicant): By express cloning, we have identified a new family of mouse receptors that associate with the signaling accessory molecule, DAP12. They have a single Ig-like domain, and their expression appears to be restricted to cells of monocyte/macrophage lineage. Thus, we have named them "MDI receptors" (for myeloid cell DAP12-associated Ig-like receptors). We have identified 3 overlapping BACs containing MDI genes and have mapped the genes to chromosome 17d. Digests of the BACs indicate that the MDI receptor family is small, probably limited to two homologous genes, MDI-1 and MDI-2. When expressed in MT2 macrophages, MDI-1 associates with endogenous DAP12 and stimulates the production of nitric oxide. Only 3 other receptors are known to associate with DAP12 in monocyte/macrophage cells. Mice deficient in Dap12 have immune alterations that appear to lie in antigen presenting cells, and humans deficient in DAP12 have neurologic and bone disorders that may reflect defects in microglial cells and osteoclasts, respectively. We therefore wish to define the functions of the MDI receptors and their ligands. We propose five specific aims: Specific Aim 1. Define the ligands for MDI-1 and MDI-2. We will create soluble MDI-1 and MDI-2 receptors to identify ligands on other cells or, if indicated, from serum or other sources, We will seek to identify the ligands either as known proteins or, if they are unknown, to clone the cDNA(s) for the ligand(s). Specific Aim 2. Define the extent of the MDI receptor family. We will further probe monocyte/macrophage cDNA libraries to seek additional members of the MDI family. Second we will map and sequence the MDI genes that are encoded in the three BACs that hybridize to MDI transcripts. Specific Aim 3. Define the range of expression of MDI receptors. We will produce monoclonal antibodies against individual MDI receptors, and we will use these to assess the surface expression of MDI on different cell types and at different stages of cell activation. Specific Aim 4. Define the cellular responses to ligation of MDI. We will study both receptor-transfected cells and freshly prepared cells. Our studies will be guided by our own results and by the phenotype of DAP12-deficient mice and humans. They include a collaboration with Dr. Jason Cyster regarding chemokines and chemokine receptors. Specific Aim 5. Define the phenotype of MDI-/- mice. In collaboration with Nigel Killeen, we will create mice deficient in both MDI-1 and MDI-2. Again, these studies will be guided by our own results as well as the phenotype of DAP12-deficient mice and humans.

描述（由申请人提供）：通过明确克隆，我们已经确定了 与信号配件相关的新型小鼠受体家族 分子，DAP12。他们有一个类似Ig的域，他们的表达 似乎仅限于单核细胞/巨噬细胞谱系的细胞。因此，我们有 将它们命名为“ MDI受体”（用于髓样细胞DAP12相关的Ig样 受体）。我们已经确定了3个包含MDI基因的重叠BAC，并且具有 将基因映射到17d染色体。 BAC的消化表明MDI 受体家族很小，可能仅限于两个同源基因，MDI-1和 MDI-2。当在MT2巨噬细胞中表达时，MDI-1与内源性相关 DAP12并刺激一氧化氮的产生。只有3个其他受体是 已知可以与单核细胞/巨噬细胞中的DAP12相关。小鼠缺乏 DAP12具有免疫改变，似乎位于抗原呈现细胞中， 人类缺乏DAP12的人具有神经系统疾病，可能 分别反映小胶质细胞和破骨细胞的缺陷。因此，我们 希望定义MDI受体及其配体的功能。我们建议 五个具体目标： 具体目标1。定义MDI-1和MDI-2的配体。我们将创建可溶性 MDI-1和MDI-2受体，以鉴定其他细胞上的配体，或者（如果指示） 从血清或其他来源，我们将寻求将配体确定为 已知的蛋白质，或者，如果未知，则可以克隆配体的cDNA（S）。 具体目标2。定义MDI受体家族的程度。我们将进一步 探针单核细胞/巨噬细胞cDNA文库寻求MDI的其他成员 家庭。其次，我们将绘制并序列编码的MDI基因 三个与MDI转录本杂交的BAC。 特定目标3。定义MDI受体的表达范围。我们将 对单个MDI受体产生单克隆抗体，我们将使用 这些评估MDI在不同细胞类型和at的表面表达 细胞激活的不同阶段。 特定目的4。定义对MDI连接的细胞反应。我们将学习 受体转染的细胞和新鲜制备的细胞均两者。我们的研究将是 以我们自己的结果和DAP12缺陷小鼠的表型为指导， 人类。其中包括与Jason Cyster博士关于趋化因子的合作 和趋化因子受体。 具体目标5。定义MDI - / - 小鼠的表型。与 奈杰尔·基林（Nigel Killeen），我们将创建缺乏MDI-1和MDI-2的小鼠。再次， 这些研究将以我们自己的结果以及表型为指导 DAP12缺陷小鼠和人类。