Biology of a New DAP12 Associated Receptor Family

描述（由申请人提供）：通过表达克隆，我们鉴定了 一个新的小鼠受体家族，与信号附件相关， 分子，DAP 12。它们有一个Ig样结构域， 似乎仅限于单核细胞/巨噬细胞谱系的细胞。因此，我们有 将其命名为“MDI受体”（用于髓样细胞DAP 12相关的Ig样受体）。 受体）。我们已经鉴定了3个含有MDI基因的重叠BAC， 将基因定位在17 d号染色体上BAC摘要表明，MDI 受体家族很小，可能局限于两个同源基因，MDI-1和 MDI-2。当在MT 2巨噬细胞中表达时，MDI-1与内源性 DAP 12和刺激一氧化氮的产生。只有三种受体 已知与单核细胞/巨噬细胞中的DAP 12相关。缺陷的小鼠 Dap 12具有似乎存在于抗原呈递细胞中的免疫改变， 缺乏DAP 12的人患有神经和骨骼疾病， 分别反映了小胶质细胞和破骨细胞的缺陷。因此我们 希望确定MDI受体及其配体的功能。我们提出 五个具体目标： 具体目标1。定义MDI-1和MDI-2的配体。我们将创建可溶性的 MDI-1和MDI-2受体，以识别其他细胞上的配体，或者，如果需要， 从血清或其他来源，我们将寻求识别配体， 已知的蛋白质，或者如果它们是未知的，则克隆配体的cDNA。 具体目标2。定义MDI受体家族的范围。我们将进一步 探测单核细胞/巨噬细胞cDNA文库以寻找MDI的其他成员 家人其次，我们将绘制和测序MDI基因编码的基因， 三个BAC与MDI转录本杂交 具体目标3。定义MDI受体的表达范围。我们将 生产针对个别MDI受体的单克隆抗体，我们将使用 这些以评估MDI在不同细胞类型上的表面表达， 细胞激活的不同阶段。 具体目标4。定义细胞对MDI连接的反应。我们将研究 受体转染的细胞和新鲜制备的细胞。我们的研究将是 根据我们自己的结果和DAP 12缺陷小鼠的表型， 人类其中包括与Jason Cyster博士在趋化因子方面的合作 和趋化因子受体。 具体目标5.定义MDI-/-小鼠的表型。协同 奈杰尔基林，我们将制造MDI-1和MDI-2缺陷的小鼠。我再次重申， 这些研究将由我们自己的结果以及 DAP 12缺陷小鼠和人类。

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.