RECEPTOR-MEDIATED ENDOCYTOSIS OF EPIDERMAL GROWTH-FACTOR BY HEPATOCYTES IN THE PERFUSED-RAT-LIVER - LIGAND AND RECEPTOR DYNAMICS
RECEPTOR-MEDIATED ENDOCYTOSIS OF EPIDERMAL GROWTH-FACTOR BY HEPATOCYTES IN THE PERFUSED-RAT-LIVER - LIGAND AND RECEPTOR DYNAMICS
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DOI：
10.1083/jcb.98.6.2148
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发表时间：
1984-01-01
影响因子：
7.8
通讯作者：
HUBBARD, AL
中科院分区：
文献类型：
--
作者：
DUNN, WA;HUBBARD, AL
利用生化和形态学技术证明，肝细胞通过受体介导的过程结合、内化和降解大量的小鼠表皮生长因子(FCG)。在配基暴露之前，.apprx。每个细胞可检测到30万个高亲和力受体，没有潜伏期，并与传统的质膜标志物共存。辣根过氧化物酶标记的EGF细胞化学定位显示，受体分布于肝细胞的整个肝窦和肝细胞侧面。当饱和浓度的EGF以35度通过肝脏灌流时。C，配体清除是两相的，快速的第一阶段为每细胞20,000个分子/分钟，在15-20分钟时急剧变化为较慢的第二阶段，每细胞2500个分子/分钟。在摄取的主要阶段，.apprx。25万个EGF分子和80%的功能受体内化到内吞囊泡中，这些囊泡可以从蔗糖梯度上的质膜和溶酶体的酶标记物中分离出来。在EGF-HRP内化2-25分钟后，细胞化学显示了配体途径，并观察到从外围的内吞体到高尔基体-溶酶体区的快速转运(T1/2.simeq)。7分钟)。至少20min内未检测到125I-EGF的降解。在加入EGF后30min内，EGF达到稳定状态，持续4h，EGF清除速率等于配体降解速率(2500分子/分钟/细胞)；内体中保持恒定的未降解配体池；可访问(即细胞表面)受体的数量保持在初始值的20%。在4h时，肝细胞内化和降解的EGF分别是初始受体数量的3倍和2.3倍，即使在放线菌胺存在的情况下也是如此，并且没有实质性的受体损失。所有这些结果表明，EGF受体是内化的，其从细胞内部位循环到表面的速度取决于配体和/或受体进入溶酶体的速度。
Biochemical and morphological techniques were used to demonstrate that hepatocytes in the perfused liver bind, internalize, and degrade substantial amounts of murine epidermal growth factor (FCG) via a receptor-mediated process. Before ligand exposure, .apprx. 300,000 high-affinity receptors were detectable per cell, displayed no latency, and codistributed with conventional plasma membrane markers. Cytochemical localization using EGF coupled to horseradish peroxidase (EGF-HRP) revealed that the receptors were distributed along the entire sinusoidal and lateral surfaces of hepatocytes. When saturating concentrations of EGF were perfused through a liver at 35.degree. C, ligand clearance was biphasic with a rapid primary phase of 20,000 molecules/min per cell that dramatically changed at 15-20 min to a slower secondary phase of 2500 molecules/min per cell. During the primary phase of uptake, .apprx. 250,000 molecules of EGF and 80% of the total functional receptors were internalized into endocytic vesicles which could be separated from enzyme markers for plasma membranes and lysosomes on sucrose gradients. The ligand pathway was visualized cytochemically 2-25 min after EGF-HRP internalization and a rapid transport from endosomes at the periphery to those in the Golgi apparatus-lysosome region was observed (t1/2 .simeq. 7 min). No 125I-EGF degradation was detected for at least 20 min. Within 30 min after EGF addition, a steady state was reached which lasted up to 4 h such that the rate of EGF clearance equalled the rate of ligand degradation (2500 molecules/min per cell); a constant pool of undegraded ligand was maintained in endosomes; and the number of accessible (i.e., cell surface) receptors remained constant at 20% of initial values. By 4 h hepatocytes had internalized and degraded 3 and 2.3 times more EGF, respectively, than the initial number of available receptors, even in the presence of cycloheximide and without substantial loss of receptors. All of these results suggest that EGF receptors are internalized and that their rate of recycling to the surface from intracellular sites is governed by the rate of entry of ligand and/or receptor into lysosomes.