CHEMISTRY OF TUMORICIDAL MACROPHAGE SURFACE ANTIGENS

我们将表征配基的特异性，配基结合部位， 配体亲和力的调节和细胞骨架的结合 白细胞黏附受体Mac-L和p150、95。麦克-L既是 补体受体和细胞黏附受体；p150，95 是一种表达在髓系细胞上的密切相关的黏附受体 大量存在于毛状白血病细胞上。遗传缺陷 这些受体导致反复的细菌感染，这些细菌感染 通常在童年时是致命的。这些受体的阿尔法亚单位 控制配基的特异性，我们假设串联重复 含有可能的二价阳离子结合部位的是配体 结合部位。为了定义配体结合位点，我们将切割和 粘贴α亚基基因克隆制备Mac-L×LFA-L和 P150，95 x LFA-Lα亚基，将使用 逆转录病毒载体与共同的β亚基相关 T淋巴瘤细胞表面。MAb表位将本地化 并与单抗对功能的影响相关。在中绑定到IC3b 溶液或在E上；与新的细胞配体结合 溶液、人造平面膜和完好的细胞上 定义特定于不同配体的结合位点，以及 这些网站对亲和力的贡献。表达方式相似 截断的阿尔法亚单位将被用来定义 跨膜区和细胞质结构域在调节细胞亲和力中的作用 配体与细胞骨架的结合。我们假设细胞 动态调节配体的黏附受体亲和力。这 将在中性粒细胞上进行天然分子测试，并进行 表达在T淋巴瘤上的重组杂合分子和截短分子 单体IC3b与细胞表面的Scatchard结合 配体，在用fMLP或佛波酯刺激后。 细胞骨架结合将由洗涤剂平行测量 增溶作用。在上述研究的同时，新的配体 Mac-L在中性粒细胞聚集中识别的分子和 对内皮细胞的黏附将用单抗来定义。配基 分子的特征和初级序列将在 蛋白质和cdna水平。识别序列，假设为 要像RGD一样，将被定义。纯天然的和重组的 分泌的分子将被用于推动配体结合 学习。FMLP可调节的Mac-2的细胞内存储位置 L中性粒细胞和S小鼠MAE-L的cDNA克隆也将 特色化的。

We will characterize the ligand specificity, ligand binding sites, regulation of affinity for ligand, and cytoskeletal association of the leukocyte adhesion receptors Mac-l and p150,95. Mac-l is both a complement receptor (CR3) and a cell adhesion receptor; p150,95 is a closely related adhesion receptor expressed on myeloid cells and abundantly on hairy leukemia cells. Genetic deficiency of these receptors results in recurring bacterial infections which are often fatal in childhood. The alpha subunits of these receptors control ligand specificity, and we hypothesize that tandem repeats which contain putative divalent cation binding sites are the ligand binding sites. To define ligand binding sites, we will cut and paste alpha subunit cDNA clones to prepare hybrid Mac-l x LFA-l and p150,95 x LFA-l alpha subunits which will be expressed using retroviral vectors in association with the common beta subunit on the surface of T lymphoma cells. MAb epitopes will be localized and correlated with MAb effect on function. Binding to iC3b in solution or on E; and binding to novel cellular ligands in solution, artificial planar membranes, and on intact cells will define the binding sites specific for different ligands, and the contribution of these sites to affinity. Similarly expressed truncated alpha subunits will be used to define the importance of transmembrane and cytoplasmic domains in regulating affinity for ligand and cytoskeletal association. We hypothesize that cells dynamically regulate adhesion receptor affinity for ligand. This will be tested for native molecules on neutrophils and for recombinant hybrid and truncated molecules expressed on T lymphoma cells by Scatchard binding of monomeric iC3b and cell surface ligands, after stimulation with fMLP or phorbol esters. Cytoskeletal association will be measured in parallel by detergent solubilization. In parallel with the above studies, novel ligand molecules recognized by Mac-l in neutrophil aggregation and adhesion to endothelial cells will be defined with MAb. Ligand molecules will be characterized and primary sequence defined at the protein and cDNA levels. Recognition sequences, hypothesized to be RGD-like, will be defined. Purified natural and recombinant secreted molecules will be used in the shove ligand binding studies. The intracellular storage site of fMLP-regulatable Mac- l in neutrophils and s murine Mae-l cDNA clone will also be characterized.