CYTOSKELETON-MEMBRANE INTERACTIONS--ANTISERUM INDUCED C

在本实验室以前的工作中， 120-160千道尔顿被确定为参与细胞基质 在哺乳动物成纤维细胞和上皮细胞中的粘附。 更 最近，我们获得了一种单克隆抗体（CSAT Mab）， 对抗破坏细胞-基质粘附的鸡成肌细胞膜 并且更微妙地干扰成纤维细胞的粘附。 CSAT 单克隆抗体识别120-160的3种膜糖蛋白的复合物 千道尔顿（CSAT ag）。 在免疫荧光研究中，CSAT ag复合物 定位于主导片层，包含应力纤维的肌动蛋白部分， 以及在良好铺展的成纤维细胞中的应力纤维末端的周边。 在最后一个位置，它与纤连蛋白共定位。 这些糖蛋白 因此，它们所处的位置与它们发挥直接作用的位置相一致， 细胞-基质粘附。 本申请中提出的实验是 设计用于：（1）生产针对每个成员的单克隆抗体， 120-160千道尔顿的细胞-基质粘附糖蛋白复合物;（2） 将糖蛋白复合物的成员相对于一个 另一个在电镜水平上位于表面膜中;（3） 定位这些糖蛋白， 细胞外基质相关成分和细胞外基质相关 在电子显微镜水平上的成分;（4）研究 细胞骨架、表面膜和 细胞外基质在粘附位点丰富的CSAT ag使用快速 冷冻深蚀刻技术。 这些研究将在 控制成纤维细胞和成肌细胞，这些细胞在扰动后 它们的粘合性能。 抗体也将用于研究 CSAT ag在胚胎细胞迁移中的作用。 单特异 抗哺乳动物细胞120-160千道尔顿糖蛋白的抗体 也将作出，以研究这些粘附的组织 粘附性黑素瘤细胞中的糖蛋白及其非粘附性变体 细胞 这一系列实验将大大增加我们的 了解跨膜信号传导机制， 正常、胚胎和恶性细胞的粘附特性。 （一）

In previous work from this laboratory, integral membrane glycoproteins of 120-160 kilodaltons were identified as being involved in cell-substratum adhesion in both mammalian fibroblasts and epithelial cells. More recently, we have been provided with a monoclonal antibody (CSAT Mab) raised against chick myoblast membranes which disrupts cell-matrix adhesion of myoblasts and more subtly perturbs the adhesion of fibroblasts. CSAT Mab recognizes a complex of 3 integral membrane glycoproteins of 120-160 kilodaltons (CSAT ag). In immunofluorescence studies, the CSAT ag complex localizes to leading lamellae, portions of actin containing stress fibers, and to the periphery of stress fiber termini in well spread fibroblasts. In the last location it colocalizes with fibronectin. These glycoproteins are thus located in positions consistent with their playing a direct role in cell-matrix adhesion. The experiments proposed in this application are designed to: (1) produce monoclonal antibodies specific for each member of the 120-160 kilodaltons cell-substratum adhesion glycoprotein complex; (2) localize the members of the glycoprotein complex with respect to one another in the surface membrane at the electron microscope level; (3) localize these glycoproteins with respect to potentially relevant cytoskeletal-association components and extracellular matrix-associated components at the electron microscope level; and (4) study the ultrastructural relationship of the cytoskeleton, surface membrane and extracellular matrix at adhesion sites rich in the CSAT ag using rapid freezing deep-etch techniques. These studies will be carried out on control fibroblasts and myoblasts, and these cells following perturbation of their adhesive properties. Antibodies will also be used to study the role of the CSAT ag in the migration of embryonic cells. Monospecific antibodies against the 120-160 kilodaltons glycoproteins of mammalian cells will also be made in order to study the organization of these adhesion glycoproteins in adherent melanoma cells and a nonadherent variant of these cells. This body of experiments should increase substantially our understanding of the transmembrane signalling mechanisms which determine the adhesive properties of normal, embryonic and malignant cells. (A)