AQUAPORIN-3 WATER CHANNEL LOCALIZATION AND REGULATION IN RAT-KIDNEY
AQUAPORIN-3 WATER CHANNEL LOCALIZATION AND REGULATION IN RAT-KIDNEY
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DOI：
10.1152/ajprenal.1995.269.5.f663
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发表时间：
1995-11-01
期刊：
影响因子：
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通讯作者：
KNEPPER, MA
中科院分区：
文献类型：
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作者：
ECELBARGER, CA;TERRIS, J;KNEPPER, MA
水通道蛋白是在包括肾脏在内的一些水运输组织中表达的水通道家族。我们使用一种多肽衍生的亲和纯化的水通道蛋白-3 （AQP-3）多克隆抗体来研究其在肾脏中的定位和调控。免疫印迹实验显示其在肾皮质和髓质均有表达，其中以内髓质基部表达最多。使用逐渐提高的离心速度对膜进行亚细胞分离，发现AQP-3主要存在于4,000和17,000 g的颗粒中，与AQP-2相反，在含有小细胞内囊泡的高速（200,000 g）颗粒中几乎不存在AQP-2。免疫细胞化学和免疫荧光研究显示，标记仅限于皮质、外髓和内髓收集管。在收集管内，主细胞被标记，而插层细胞未被标记。与先前的免疫荧光研究一致(石桥、佐佐木、福岛、内田、库原、齐藤、古川、中岛、山口、Gojobori和丸莫。Proc。国家的。学会科学。美国科学（英文版）91（英文版）；1994；T. Ma， A. Frigeri， H. Hasegawa， A. S. Verkman。生物。化学，269:21845-21849,1994)，标记仅限于基侧域。在超薄冷冻切片中使用免疫金技术的免疫电子显微镜显示了基底外侧质膜的主要标记。与先前AQP-2的研究结果相反，只有有限的AQP-3标记细胞内囊泡，这表明该水通道不受囊泡运输的急性调节。免疫印迹研究显示，大鼠口渴48小时后，髓质内AQP-3蛋白的含量大约增加了一倍。这些研究与AQP-3在收集管上皮渗透驱动的吸水中的作用一致，并表明AQP-3的表达是长期调节的。
The aquaporins are a family of water channels expressed in several water-transporting tissues, including the kidney. We have used a peptide-derived, affinity-purified polyclonal antibody to aquaporin-3 (AQP-3) to investigate its localization and regulation in the kidney. Immunoblotting experiments showed expression in both renal cortex and medulla, with greatest expression in the base of the inner medulla. Subcellular fractionation of membranes, using progressively higher centrifugation speeds, revealed that AQP-3 is present predominantly in the 4,000 and 17,000 g pellets and, in contrast to AQP-2, is virtually absent in the high-speed (200,000 g) pellet that contains small intracellular vesicles. Immunocytochemistry and immunofluorescence studies revealed that labeling is restricted to the cortical, outer medullary, and inner medullary collecting ducts. Within the collecting duct, principal cells were labeled, whereas intercalated cells were unlabeled. Consistent with previous immunofluorescence studies (K. Ishibashi, S. Sasaki, K. Fushimi, S. Uchida, M. Kuwahara, H. Saito, T. Furukawa, K. Nakajima, Y. Yamaguchi, T. Gojobori, and F. Marumo. Proc. Natl. Acad. Sci. USA 91: 6269-6273, 1994; T. Ma, A. Frigeri, H. Hasegawa, and A. S. Verkman. J. Biol. Chem. 269: 21845-21849, 1994), the labeling was confined to the basolateral domain. Immunoelectron microscopy, using the immunogold technique in ultrathin cryosections, demonstrated a predominant labeling of the basolateral plasma membranes. In contrast to previous findings with AQP-2, there was only limited AQP-3 labeling of intracellular vesicles, suggesting that this water channel is not regulated acutely through vesicular trafficking. Immunoblotting studies revealed that thirsting of rats for 48 h approximately doubled the amount of AQP-3 protein in the inner medulla. These studies are consistent with a role for AQP-3 in osmotically driven water absorption across the collecting duct epithelium and suggest that the expression of AQP-3 is regulated on a long-term basis.