Wall D A, Patel S
Dev Biol. 1987 Jan;119(1):275-89. doi: 10.1016/0012-1606(87)90229-6.
A combination of electron microscopic tracers and subcellular fractionation has been used to examine the endocytic pathway of the yolk protein precursor, vitellogenin (VG), in Xenopus oocytes. VG was adsorbed to colloidal gold, and the organelles traversed by newly internalized ligand were examined at various time intervals after endocytosis. VG-Au enters oocytes via coated pits and vesicles and then appears rapidly in tubular endosomes and multivesicular bodies (MVBs). MVBs play a central role in VG processing for storage; the large majority of newly internalized VG enters this compartment, remaining there for up to several hours. Condensation of VG into crystalline bodies begins in MVBs, and continues with growth of the crystals until typical platelets are formed. When oocytes are exposed to high [VG], MVBs containing large amounts of internalized VG are morphologically indistinguishable from the primordial yolk platelets described earlier (Dumont, 1978). The use of VG-Au particles of two sizes demonstrates that gold particles in early MVBs were generally associated with the limiting membrane of these organelles, while older MVB compartments have gold particles well separated from the limiting membranes, suggesting that dissociation of VG from its receptor occurs in this compartment. Newly internalized ligand preferentially forms a new MVB, rather than fusing and mixing with previously formed MVBs. Progressive yolk protein condensation gradually transforms MVBs into yolk platelets over a period of several hours. Analysis of 125I-VG-Au behavior after sucrose gradient fractionation of oocytes allowed correlation of biochemical compartments with those observed in the electron microscope. MVBs containing yolk in progressive stages of condensation were found at densities from 1.16 up to 1.21 g/cc. The final, rate-limiting step in VG transport is a shift of ligand from light (1.21 g/cc) to heavy (1.23 g/cc) platelet compartments (Wall and Meleka, 1985). The morphological correlate of this process is movement of VG-Au from small (less than 3-4 microns diameter) to large (greater than 4 microns diameter) platelets.
电子显微镜示踪剂与亚细胞分级分离相结合,已被用于研究非洲爪蟾卵母细胞中卵黄蛋白前体——卵黄原蛋白(VG)的内吞途径。将VG吸附到胶体金上,并在胞吞作用后的不同时间间隔检查新内化配体所穿过的细胞器。VG-金通过被膜小窝和小泡进入卵母细胞,然后迅速出现在管状内体和多囊泡体(MVBs)中。MVBs在用于储存的VG加工过程中起核心作用;绝大多数新内化的VG进入这个区室,并在那里停留长达数小时。VG凝聚成晶体开始于MVBs,并随着晶体生长持续进行,直到形成典型的血小板。当卵母细胞暴露于高浓度[VG]时,含有大量内化VG的MVBs在形态上与之前描述的原始卵黄血小板无法区分(Dumont,1978)。使用两种大小的VG-金颗粒表明,早期MVBs中的金颗粒通常与这些细胞器的界膜相关,而较老的MVB区室中的金颗粒与界膜分隔良好,这表明VG与其受体在这个区室中发生解离。新内化的配体优先形成新的MVB,而不是与先前形成的MVB融合并混合。在数小时内,卵黄蛋白的逐步凝聚逐渐将MVBs转化为卵黄血小板。对卵母细胞进行蔗糖梯度分级分离后对125I-VG-金行为的分析,使生化区室与电子显微镜下观察到的区室相关联。在密度为1.16至1.21 g/cc的区域发现了处于逐步凝聚阶段且含有卵黄的MVBs。VG运输的最后限速步骤是配体从轻(1.21 g/cc)血小板区室向重(1.23 g/cc)血小板区室的转移(Wall和Meleka,1985)。这个过程的形态学对应是VG-金从小(直径小于3 - 4微米)血小板向大(直径大于4微米)血小板的移动。