CHARACTERIZATION OF MAST CELL INTEGRINS

拟议研究的目标将是了解的作用， 新型整联蛋白在肥大细胞生物学中起作用。 成熟肥大细胞 定位于不同的组织以完成不同的功能 免疫反应的抗原特异性IgE分支。 的机制 肥大细胞知道它的最终目的地在哪里， 尽管过去的共识是这些细胞定位于 响应特定细胞因子/淋巴因子需求， 受体的 我们发现成熟的粘膜肥大细胞表达一种新的 β整合素，我们假设，这种整合素，与 一个或多个α链将肥大细胞定位于特定组织。 该假设的证明是本申请中提出的研究。 首先，这种β整联蛋白的细胞表面表达必须是 确认并鉴定与其相关的α链。 的 这些整合素链的表达的调节将在 肥大细胞分化的不同条件。 细胞表面表达 将通过使用预先存在的抗体来监测这些链 我们从这些整合素分子中提取的特异性分子。 的 新β和/或α整联蛋白链的鉴定和分离 将通过PCR克隆这些基因来实现， 保守编码序列的扩增，低严格性杂交 与其他整联蛋白链cDNA或新型整联蛋白的肽测序 链被鉴定为已知整联蛋白链的伴侣。 归巢 这些复合物的功能将通过阻断抗体来证实， 反义构建体，或通过 将β和α链编码序列转染到细胞中 通常不会表达出来。 归巢分析将在 使用正常和肥大细胞缺陷品系的小鼠进行体外和体内试验。 这种归巢功能的最终证明将在老鼠的一代中 缺乏β和/或α链的表达， 与缺陷基因序列重组。 利用这种基因， 细胞生物学和生物化学实验方法，这些方法的作用 将测定肥大细胞生物学中的整联蛋白复合物。

The goal of the proposed research will be to understand the role that a novel integrin plays in the biology of the mast cell. Mature mast cells localize to different tissues to accomplish different functions within the antigen specific, IgE, arm of the immune response. The mechanism by which a mast cell knows where its eventual destination is has not been explored although the past consensus has been that these cells localize in response to specific cytokine/lymphokine requirements not a homing receptor. We have found that mature mucosal mast cells express a novel beta integrin and we hypothesize that this integrin, in association with one or more alpha chains, localizes mast cells to specific tissues. The proof of the hypothesis is the research proposed in this application. First, the cell surface expression of this beta integrin must be confirmed and the alpha chains it is associated with be identified. The modulation of expression of these integrin chains will be studied under varying conditions of mast cell differentiation. Cell surface expression of these chains will be monitored via the use of pre-existing antibodies those we derive specific for these integrin molecules. The identification and isolation of novel beta and/or alpha integrin chains will be accomplished via the cloning of such genes either through PCR amplification of conserved coding sequences, low stringency hybridization with other integrin chain cDNA's, or peptide sequencing of novel integrin chains identified as partners to known integrin chains. The homing function of these complexes will be confirmed via blocking antibody, antisense constructs, or functional transfer of homing activity via transfection of the beta and alpha chain coding sequences into a cell that does not normally express them. Homing anayses will be done both in vitro and in vivo using normal and mast cell deficient strains of mice. The final proof of this homing function will be in the generation of mice lacking the beta and/or alpha chain expression through homologous recombination with a defective gene sequence. Using this mix of genetic, cell biology and biochemical experimental approaches, the roles of these integrin complexes in the biology of the mast cell will be determined.