CHARACTERIZATION OF MAST CELL INTEGRINS

拟议研究的目标将是了解一个 新的整合素在肥大细胞的生物学中发挥作用。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臂。这个机制是通过 主信元知道其最终目的地未到达的 尽管过去的共识是这些细胞定位于 响应特定的细胞因子/淋巴因子要求，而不是归巢 受体。我们发现成熟的粘膜肥大细胞表达一种新的 并且我们假设这个整合素，与 一个或多个阿尔法链，将肥大细胞定位于特定的组织。 这一假设的证明是本申请中提出的研究。 首先，这种β整合素的细胞表面表达必须是 确认并识别与其相关联的阿尔法链。这个 这些整合素链的表达调控将在以下方面进行研究 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 我们为这些整合素分子所特有的那些。这个 新的β和/或α整合素链的鉴定和分离 将通过通过聚合酶链式反应克隆这些基因来实现 扩增保守编码序列，低严格率杂交 与其他整合素链cDNAs，或新整合素的肽序列 确定为已知整合素链的伙伴的链。归宿 这些复合体的功能将通过封闭抗体来确认， 反义构建，或功能转移的归巢活动通过 将β和阿尔法链编码序列导入细胞 这通常不会表达他们的意思。归宿分析将在两个版本中完成 体外和体内使用正常和肥大细胞缺陷品系的小鼠。 这种归巢功能的最终证据将是小鼠的生成 缺乏β和/或阿尔法链表达的同源 与有缺陷的基因序列重组。利用这种混合的基因， 细胞生物学和生化实验方法，这些方法的作用 肥大细胞生物学中的整合素复合体将被确定。

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.