Ferland L H, Djiane J, Houdebine L M, Kelly P A
Endocrinology. 1984 Nov;115(5):1842-9. doi: 10.1210/endo-115-5-1842.
PRL receptors have been previously identified in purified rat liver plasma membrane and Golgi vesicle preparations. In this study, we report on PRL receptors located in highly purified lysosome preparations. These lysosomal PRL receptors were characterized using Scatchard analysis and compared to other intracellular and cell surface receptors. We have identified two classes of lysosomes. Lighter lysosome-like vesicles, which are greatly enriched in acid phosphatase activity (the marker enzyme of lysosomes), contain a great deal of binding activity. This PRL binding was only slightly increased by pretreatment of animals with the lysosomotropic agent chloroquine. In contrast, mature lysosomes showed very little binding activity in control animals, but chloroquine treatment increased binding 7- to 8-fold in these mature lysosomes. We suggest that the lysosome-like structures are immature lysosomes (namely prelysosomes) toward which the hormone-receptor complex is internalized: they appear to bear little proteolytic activity. These structures could play a role in PRL receptor recycling. Lysosomal PRL receptors showed curvilinear Scatchard plots, in contrast to plasma membrane and Golgi counterparts, which were linear over the same range of hormone concentrations. The high affinity site in lysosomes had a Kd comparable to the cell surface and Golgi receptors. The number of binding sites per mg protein in prelysosomes and lysosomes was 3 times greater than that in the homogenate, but Golgi preparations were 3 times as rich as lysosomes. The great number of PRL receptors in prelysosomes could be attributed, in large part, to the low affinity sites. The internalization of PRL into rat liver was examined after in vivo injection of [125I]iodoovine PRL. The labeled hormone was found initially in the plasma membrane fraction, after which it localized preferentially in the Golgi fraction, with maximum incorporation 15 min postinjection. Substantial radioactivity was observed in both classes of lysosomes (L-1 and L-2). In contrast to the Golgi fraction, maximum incorporation of [125I]iodoovine PRL in lysosomes occurred at 30 min. This suggests either that during internalization, PRL first reaches Golgi elements and is then transferred to the lysosomal compartment, or that there are two independent pathways of internalization, one rapid toward the Golgi complex (may be a path of receptor recycling) and the other toward lysosomes (probably leading to receptor degradation).
催乳素（PRL）受体先前已在纯化的大鼠肝细胞膜和高尔基体囊泡制剂中被鉴定出来。在本研究中，我们报道了位于高度纯化的溶酶体制剂中的PRL受体。使用Scatchard分析对这些溶酶体PRL受体进行了表征，并与其他细胞内和细胞表面受体进行了比较。我们鉴定出了两类溶酶体。较轻的类溶酶体囊泡，其酸性磷酸酶活性（溶酶体的标志性酶）大大富集，具有大量的结合活性。用溶酶体促渗剂氯喹预处理动物后，这种PRL结合仅略有增加。相比之下，成熟溶酶体在对照动物中显示出非常低的结合活性，但氯喹处理使这些成熟溶酶体中的结合增加了7至8倍。我们认为类溶酶体结构是未成熟的溶酶体（即前溶酶体），激素 - 受体复合物被内化到其中：它们似乎几乎没有蛋白水解活性。这些结构可能在PRL受体循环中起作用。与细胞膜和高尔基体对应物不同，溶酶体PRL受体显示出曲线型的Scatchard图，而细胞膜和高尔基体对应物在相同激素浓度范围内呈线性。溶酶体中的高亲和力位点的解离常数（Kd）与细胞表面和高尔基体受体相当。前溶酶体和溶酶体中每毫克蛋白质的结合位点数比匀浆中的多3倍，但高尔基体制剂的丰富程度是溶酶体的3倍。前溶酶体中大量的PRL受体在很大程度上可归因于低亲和力位点。在体内注射[125I]碘代羊催乳素后，检测了PRL在大鼠肝脏中的内化情况。标记的激素最初在细胞膜部分被发现，之后它优先定位于高尔基体部分，注射后15分钟掺入量达到最大。在两类溶酶体（L - 1和L - 2）中均观察到大量放射性。与高尔基体部分不同，溶酶体中[125I]碘代羊催乳素的最大掺入量在30分钟时出现。这表明在内化过程中，PRL要么首先到达高尔基体成分，然后转移到溶酶体区室，要么存在两条独立的内化途径，一条快速通向高尔基体复合体（可能是受体循环途径），另一条通向溶酶体（可能导致受体降解）。
