Brown D, Sabolić I
Renal Unit, Massachusetts General Hospital, Boston.
J Cell Sci Suppl. 1993;17:49-59. doi: 10.1242/jcs.1993.supplement_17.8.
The plasma membrane composition of virtually all eukaryotic cells is maintained and continually modified by the recycling of specific protein and lipid components. In the kidney collecting duct, urinary acidification and urinary concentration are physiologically regulated at the cellular level by the shuttling of proton pumps and water channels between intracellular vesicles and the plasma membrane of highly specialized cell types. In the intercalated cell, hydrogen ion secretion into the urine is modulated by the recycling of vesicles carrying a proton pumping ATPase to and from the plasma membrane. In the principal cell, the antidiuretic hormone, vasopressin, induces the insertion of vesicles that contain proteinaceous water channels into the apical cell membrane, thus increasing the permeability to water of the epithelial layer. In both cell types, 'coated' carrier vesicles are involved in this process, but whereas clathrin-coated vesicles are involved in the endocytotic phase of water channel recycling, the transporting vesicles in intercalated cells are coated with the cytoplasmic domains of the proton pumping ATPase. By a combination of morphological and functional techniques using FITC-dextran as an endosomal marker, we have shown that recycling endosomes from intercalated cells are acidifying vesicles but that they do not contain water channels. In contrast, principal cell vesicles that recycle water channels do not acidify their lumens in response to ATP. These non-acidic vesicles lack functionally important subunits of the vacuolar proton ATPase, including the 16 kDa proteolipid that forms the transmembrane proton pore. Because these endosomes are directly derived via clathrin-mediated endocytosis, our results indicate that endocytotic clathrin-coated vesicles are non-acidic compartments in principal cells. In contrast, recycling vesicles in intercalated cells contain large numbers of proton pumps, arranged in hexagonally packed arrays on the vesicle membrane. These pumps are inserted into the apical plasma membrane of A-type (acid-secreting) intercalated cells, and the basolateral plasma membrane of B-type (bicarbonate-secreting) cells in the collecting duct. Both apical and basolateral targeting of H(+)-ATPase-containing vesicles in these cells may be directed by microtubules, because polarized insertion of the pump into both membrane domains is disrupted by microtubule depolymerizing agents. However, the basolateral localization of other transporting proteins in intercalated cells, including the band 3-like anion exchanger and facilitated glucose transporters, is not affected by microtubule disruption.
几乎所有真核细胞的质膜组成都是通过特定蛋白质和脂质成分的循环利用来维持并不断修饰的。在肾集合管中，尿液酸化和尿液浓缩在细胞水平上通过质子泵和水通道在细胞内囊泡与高度特化细胞类型的质膜之间穿梭来进行生理调节。在闰细胞中，向尿液中分泌氢离子是通过携带质子泵ATP酶的囊泡在质膜的往返循环来调节的。在主细胞中，抗利尿激素血管加压素会促使含有蛋白质性水通道的囊泡插入顶端细胞膜，从而增加上皮层对水的通透性。在这两种细胞类型中，“有被”载体囊泡都参与了这个过程，不过网格蛋白包被囊泡参与水通道循环的内吞阶段，而闰细胞中的转运囊泡则被质子泵ATP酶的细胞质结构域包被。通过使用FITC - 葡聚糖作为内体标记物的形态学和功能技术相结合，我们发现闰细胞的循环内体是酸化囊泡，但它们不含水通道。相比之下，回收水通道的主细胞囊泡不会因ATP而使其内腔酸化。这些非酸性囊泡缺乏液泡质子ATP酶的功能重要亚基，包括形成跨膜质子孔的16 kDa蛋白脂质。由于这些内体是通过网格蛋白介导的内吞作用直接产生的，我们的结果表明，内吞的网格蛋白包被囊泡在主细胞中是非酸性区室。相比之下，闰细胞中的循环囊泡含有大量质子泵，在囊泡膜上以六边形排列。这些泵被插入到集合管中A型（分泌酸的）闰细胞的顶端质膜和B型（分泌碳酸氢盐的）细胞的基底外侧质膜中。这些细胞中含H(+) - ATP酶囊泡的顶端和基底外侧靶向可能都由微管引导，因为微管解聚剂会破坏泵向两个膜结构域的极化插入。然而，闰细胞中其他转运蛋白的基底外侧定位，包括带3样阴离子交换器和易化葡萄糖转运体，不受微管破坏的影响。
