Painter R G, Ginsberg M
J Cell Biol. 1982 Feb;92(2):565-73. doi: 10.1083/jcb.92.2.565.
We have measured the association of platelet surface membrane proteins with Triton X-100 (Triton)-insoluble residues in platelets surface labeled with 125I. In both concanavalin A (Con A)-stimulated and resting platelets, this fraction is composed largely of polypeptides with apparent molecular weights of 45,000, 200,000, and 250,000 which comigrate with authentic actin, myosin heavy chain, and actin binding protein, respectively, as judged by PAGE in SDS. Less than 10% of the two major 125I-labeled surface glycoproteins, GPiib and GPIII, were associated with the Triton residue in resting platelets. Within 45 s after Con A addition, 80-95% of these two glycoproteins became associated with the Triton residue and the amount of sedimentable actin doubled. No cosedimentation of GPIIb and III with the cytoskeletal protein-containing Triton residue was seen when Con A was added to a Triton extract of resting cells, indicating that the sedimentation of GPIIb and III seen in Con A-stimulated platelets was not due to precipitation of the glycoproteins by Con A after detergent lysis. Treatment of Triton extracts of Con A-stimulated platelets with DNase I (deoxyribonucleate 5'-oligonucleotidido-hydrolase [EC 3.1.4.5]) inhibited the sedimentation of actin and the two surface glycoproteins in a dose-dependent manner. This inhibition of cosedimentation was not due to an effect of DNase I on Con A-glycoprotein interactions since these two glycoproteins could be quantitatively recovered by Con A-Sepharose affinity absorption in the presence of DNase I. When the Con A bound to the Triton residue was localized ultrastructurally, it was associated with cell-sized structures containing filamentous material. In intact cells, there was simultaneous immunofluorescent coredistribution of surface-bound Con A and myosin under conditions which induced a redistribution of platelet myosin. These data suggest that Con A can, in the intact platelet, induce physical interactions between certain surface glycoproteins and the internal cytoskeleton.
我们测定了血小板表面膜蛋白与用¹²⁵I进行表面标记的血小板中不溶于 Triton X - 100（Triton）的残渣之间的关联。在伴刀豆球蛋白 A（Con A）刺激的血小板和静息血小板中，该部分主要由表观分子量分别为 45,000、200,000 和 250,000 的多肽组成，通过 SDS 中的 PAGE 判断，它们分别与真实的肌动蛋白、肌球蛋白重链和肌动蛋白结合蛋白共迁移。在静息血小板中，两种主要的¹²⁵I 标记表面糖蛋白 GPIIb 和 GPIII 与 Triton 残渣的关联不到 10%。添加 Con A 后 45 秒内，这两种糖蛋白的 80 - 95%与 Triton 残渣相关联，并且可沉降的肌动蛋白量增加了一倍。当将 Con A 添加到静息细胞的 Triton 提取物中时，未观察到 GPIIb 和 III 与含细胞骨架蛋白的 Triton 残渣的共沉降，这表明在 Con A 刺激的血小板中观察到的 GPIIb 和 III 的沉降不是由于去污剂裂解后 Con A 使糖蛋白沉淀所致。用 DNase I（脱氧核糖核酸 5'-寡核苷酸水解酶[EC 3.1.4.5]）处理 Con A 刺激的血小板的 Triton 提取物，以剂量依赖的方式抑制了肌动蛋白和两种表面糖蛋白的沉降。这种共沉降的抑制不是由于 DNase I 对 Con A - 糖蛋白相互作用的影响，因为在存在 DNase I 的情况下，这两种糖蛋白可以通过 Con A - Sepharose 亲和吸附定量回收。当超微结构定位与 Triton 残渣结合的 Con A 时，它与含有丝状物质的细胞大小的结构相关联。在完整细胞中，在诱导血小板肌球蛋白重新分布的条件下，表面结合的 Con A 和肌球蛋白同时出现免疫荧光共分布。这些数据表明，在完整的血小板中，Con A 可以诱导某些表面糖蛋白与内部细胞骨架之间的物理相互作用。
