Bozyczko D, Decker C, Muschler J, Horwitz A F
Department of Biochemistry and Biophysics, University of Pennsylvania School of Medicine, Philadelphia 19104.
Exp Cell Res. 1989 Jul;183(1):72-91. doi: 10.1016/0014-4827(89)90419-9.
Avian integrin is a complex of integral membrane glycoproteins that appears to function as a dual receptors for both intracellular cytoskeletal and extracellular matrix components. Antibodies were raised against this complex and used to (1) immunolocalize integrin on cryosections of developing and adult muscle tissue and on developing myotube cultures in vitro and (2) immunoaffinity purify integrin from various fiber-type specific muscles. Integrin localization was compared with that of its putative cytoskeletal-associated and extracellular matrix ligands, talin and vinculin and fibronectin and laminin, respectively. The goal was to identify putative sites of interaction between the muscle sarcolemma and the cytoskeleton and the extracellular matrix and to reveal any differences in the molecular composition at these sites. Integrin's distribution on the sarcolemma of early (Day 12) embryonic limb muscle was random and punctate. On late embryonic (Days 17-19) limb muscle tissue its distribution was generally uniform but with occasional increased densities at specific sites along the sarcolemma. Posthatch (greater than 3 weeks) fast twitch muscle showed a highly regionalized distribution. These regions of integrin concentration coincided with densities of acetylcholine receptors, revealed by TRITC alpha-bungarotoxin labeling, and regions of muscle-tendon interaction, identified by morphological criteria. Tissue culture studies also demonstrated integrin densities at analogous sites in vitro, e.g., acetylcholine receptor clusters and sites at which myofibrils terminate at the sarcolemma. These integrin-rich sites were also shown to be Triton X-100 insoluble and therefore presumably are linked to the cytoskeleton or extracellular matrix. The localization of integrin on developing and adult muscle tissue was compared with that of fibronectin, laminin, vinculin, and talin using double, immunofluorescently labeled cryosections. In general, integrin did not colocalize exclusively with any one of its putative ligands. In the embryo, discrete densities of both talin and vinculin were observed at the myotendinous junction, whereas integrin immunoreactivity was widely distributed on muscle, vasculature, nerve, and connective tissue with no discernible sites of increased density. Laminin was primarily associated with muscle and nerve whereas fibronectin was prominent on connective tissue. On posthatch tissue, the distributions of talin, vinculin, laminin, and fibronectin were similar to those in the embryo, whereas the distribution of integrin was restricted to specific sites. The distribution of integrin was also examined for fiber-type specific differences on adu
禽整联蛋白是一种整合膜糖蛋白复合物，似乎作为细胞内细胞骨架和细胞外基质成分的双重受体发挥作用。制备了针对这种复合物的抗体，并用于（1）在发育中和成年肌肉组织的冷冻切片以及体外培养的发育中的肌管上对整联蛋白进行免疫定位，以及（2）从各种纤维类型特异性肌肉中免疫亲和纯化整联蛋白。将整联蛋白的定位分别与其假定的细胞骨架相关和细胞外基质配体、踝蛋白和纽蛋白以及纤连蛋白和层粘连蛋白的定位进行比较。目的是确定肌肉肌膜与细胞骨架和细胞外基质之间假定的相互作用位点，并揭示这些位点在分子组成上的任何差异。整联蛋白在早期（第12天）胚胎肢体肌肉的肌膜上的分布是随机且点状的。在胚胎后期（第17 - 19天）肢体肌肉组织上，其分布通常是均匀的，但在肌膜上的特定部位偶尔会有密度增加。孵化后（大于3周）的快肌显示出高度区域化的分布。这些整联蛋白浓度区域与用TRITCα - 银环蛇毒素标记显示的乙酰胆碱受体密度以及通过形态学标准确定的肌肉 - 肌腱相互作用区域一致。组织培养研究还证明了体外类似部位的整联蛋白密度，例如乙酰胆碱受体簇以及肌原纤维在肌膜处终止的部位。这些富含整联蛋白的部位也被证明对Triton X - 100不溶，因此推测与细胞骨架或细胞外基质相连。使用双重免疫荧光标记的冷冻切片，将整联蛋白在发育中和成年肌肉组织上的定位与纤连蛋白、层粘连蛋白、纽蛋白和踝蛋白的定位进行比较。一般来说，整联蛋白并不完全与它的任何一种假定配体共定位。在胚胎中，在肌腱连接处观察到踝蛋白和纽蛋白的离散密度，而整联蛋白免疫反应性广泛分布在肌肉、脉管系统、神经和结缔组织上，没有明显的密度增加部位。层粘连蛋白主要与肌肉和神经相关，而纤连蛋白在结缔组织上突出。在孵化后组织上，踝蛋白、纽蛋白、层粘连蛋白和纤连蛋白的分布与胚胎中的相似，而整联蛋白的分布局限于特定部位。还检查了成年期整联蛋白分布的纤维类型特异性差异
