125I-thrombin binds to clustered receptors on noncoated regions of mouse embryo cell surfaces.
125I-thrombin binds to clustered receptors on noncoated regions of mouse embryo cell surfaces.
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125I-凝血酶与小鼠胚胎细胞表面非包被区域的聚集受体结合。
DOI：
10.1083/jcb.95.3.697
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发表时间：
1982
期刊：
影响因子：
--
通讯作者：
Bergmann,JS
中科院分区：
文献类型：
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作者：
Carney,DH;Bergmann,JS
我们利用电子显微镜放射自显像（EMAR）观察了1251凝血酶与其表面受体在小鼠胚胎细胞上的相互作用。放射自显像颗粒以周期性的非随机模式分布在细胞表面，表明12~凝血酶与凝血酶受体簇相关。细胞间的颗粒间距略有不同，表明细胞亚群具有不同数量的凝血酶受体。1251-凝血酶（125 ng/ml）在37℃下与ME细胞结合后，ME细胞上颗粒之间的平均距离为1.65±0.49# m，而固定细胞和12l -凝血酶在4℃下与37℃下的颗粒之间的平均距离没有显著差异。这表明凝血酶受体在结合前聚集，凝血酶受体聚集体在凝血酶结合后不会重新分布成细胞表面的大聚集体。在这三种约束条件下，每簇颗粒的数量也不会改变。因此，每个簇中被占用的受体的数量似乎是恒定的。根据平均颗粒数和间距，我们估计每个簇的直径约为400nm，包含大约550个凝血酶结合位点。这些受体簇与膜的特殊结构或涂覆区域无关。此外，在细胞内观察到的颗粒未发现与包被囊泡相关。因此，~ 251-凝血酶的聚集模式和内化都不是与受体结合并通过受体介导的内吞作用内化的分子的特征。包括低密度脂蛋白（LDL）、asialoorosomucnid、t~ z-巨球蛋白、胰岛素和表皮生长因子（EGF）在内的许多分子与细胞表面受体结合，这些受体聚集在被涂膜区域，并通过称为受体介导的内吞作用的过程被内化（1,2）。然而，目前尚不清楚这种类型的内吞作用是否在激素和生长因子的生物活性中起作用，或者它是否仅仅作为调节这些分子或其受体水平的降解途径。一些未被占用的受体在配体结合前聚集在细胞表面。在与低密度脂蛋白孵育前，将成纤维细胞置于甲醛中或冷却至4℃，似乎有很大比例的低密度脂蛋白受体预先聚集(3)。此外，高达58%的asialoorosomucoid受体可能在配体结合前聚集(4)。在这两种情况下，这些簇都与涂层凹坑相关联，以便内化。据报道，激素类分子最初也与聚集的受体结合。例如，乙酰胆碱受体似乎在肌细胞表面聚集，而不是内胆碱受体
We used electron microscope autoradiography (EMAR) to visualize the interaction of 1251-thrombin with its surface receptors on mouse embryo (ME) cells. Autoradiographic grains were spaced over the surface of cells in a periodic nonrandom pattern, indicating 12~ lthrombin association with clusters of thrombin receptors. The grain spacing varied slightly from cell to cell, indicating subpopulations of cells with different numbers of thrombin receptors. The average distance between grains on ME cells after binding 1251-thrombin (125 ng/ml) at 37 C was 1.65 _+ 0.49# m. The average distance between grains on prefixed cells and cells incubated with 12Sl-thrombin at 4 C was not significantly different from that observed at 37 C. This indicates that thrombin receptors are clustered before thrombin binding and that the thrombin receptor aggregates do not redistribute into large aggregates on the surface of cells subsequent to thrombin binding. The number of grains per cluster also does not change under these three binding conditions. Thus, the number of occupied receptors in each cluster appears to be constant. On the basis of the average grain number and spacing, we estimate that each cluster is~ 400 nm in diameter containing approximately 550 thrombin-binding sites. These receptor-clusters are not associated with specialized structures or coated regions of the membrane. Additionally, grains observed within cells were not found associated with coated vesicles. Therefore, neither the clustering patterns nor internalization of~ 251-thrombin are characteristic of molecules which bind to receptors and are internalized by receptor-mediated endocytosis.A number of molecules including low density lipoprotein (LDL), asialoorosomucnid, t~ z-macroglobulin, insulin and epidermal growth factor (EGF) bind to cell surface receptors which aggregate in coated membrane regions and are internalized by the process referred to as receptor-mediated endocytosis (1, 2). It is not clear, however, whether this type of endocytosis plays a role in the biological activity of hormones and growth factors or whether it merely serves as a degradative pathway to regulate the levels of these molecules or their receptors. Some unoccupied receptors are aggregated on the cell surface before ligand binding. Fibroblasts prefixed in formaldehyde or chilled to 4 C before incubation with LDL appear to have a large proportion of their LDL receptors preclustered (3). In addition, up to 58% of the asialoorosomucoid receptors may be clustered before ligand binding (4). In both cases, these clusters associate with coated pits for internalization. Hormonelike molecules have also been reported to initially bind to aggregated receptors. For example, acetylcholine receptors appear to be aggregated on the surface of muscle ceils prior to inner-