Yasuko Tanaka

The role of mammalian superaquaporins inside the cell.

BACKGROUND The mammalian two superaquaporins, AQP11 and AQP12, are present inside the cell and their null phenotypes in mice suggest their unusual functions. SCOPE OF REVIEW The surveyed literature on these superaquaporins and our unpublished data has been incorporated to speculate their roles. MAJOR CONCLUSIONS AQP11 and AQP12 have unique NPA boxes with a signature cysteine residue. Although some water permeability of AQP11 was demonstrated in liposomes and cultured cells, its permeability to glycerol is unknown. The function of AQP12 still remains to be clarified. AQP11 null mice develop polycystic kidneys following large intracellular vacuoles in the proximal tubule, which may be caused by ER stress or vesicle fusion failure. The role of AQP11 in the kidney and liver seems to alleviate the tissue damage and facilitate the recovery. Its expression in the sperm, thymus and brain suggests its potential roles in these organs in spite of the apparently normal null phenotype. Although AQP12 null mice appear normal, they suffer from severe pancreatitis, suggesting its role in the fusion of zymogen granules. GENERAL SIGNIFICANCE As many issues are unsolved, the clarification of the function and roles of the superaquaporin may lead to the identification of new roles of AQPs. This article is part of a Special Issue entitled Aquaporins.

Aquaporin-11 knockout mice and polycystic kidney disease animals share a common mechanism of cyst formation

Aquaporin‐11 (AQP11), a new member of the aquaporin family, is localized in the endoplasmic reticulum (ER). Aqp11−/− mice neonatally suffer from polycystic kidneys derived from the proximal tubule. Its onset is proceeded by the vacuolization of ER. However, the mechanism for the formation of vacuoles and cysts remains to be clarified. Here, we show that Aqp11−/− mice and polycystic kidney disease (PKD) animals share a common pathogenic mechanism of cyst formation. We performed microarray analyses and histochemical staining to characterize the effects of the disruption of Aqp11 on kidneys of 1‐wk‐old mice. Microarray analyses revealed that the significantly changed functional categories in Aqp11−/− mice were similar to those in PKD animals. Histochemical studies showed expression changes of 3 genes, Myc, Egfr, Egf, which are assumed to be involved in the proliferation of cystic cells in PKD. We actually confirmed the activation of cell proliferation in the proximal tubule cells with vacuolized ER. Furthermore, three genes associated with the remodeling of the extracellular matrix, Mmp12, Timpl, Tgfbl, were up‐regulated in the fibroblasts. We also demonstrated the activation of apoptosis via the ER‐stress pathway in the proximal tubule cells with vacuolized ER. These results provide new insights into the physiological roles of AQP11.—Okada, S., Misaka, T., Tanaka, Y., Matsumoto, I., Ishibashi, K., Sasaki, S., Abe, K. Aquaporin‐11 knockout mice and polycystic kidney disease animals share a common mechanism of cyst formation. FASEB J. 22, 3672–3684 (2008)

Aquaporin10 is a pseudogene in cattle and their relatives

Background Although AQP10 is mainly expressed in the human GI tract, its physiological role is unclear. In fact, we previously reported that mouse AQP10 is a pseudogene. It is possible that AQP10 is also a pseudogene in other animals. Methods Genome databases were searched for AQP10 orthologs and the genomic DNA of each candidate pseudogene was sequenced to confirm its mutations. The expression of the AQP10 mRNA was examined by RT-PCR in the small intestine where human AQP10 is highly expressed. Results The genomic database of some mammals had insertions and deletions in the exons of the AQP10 gene, including cattle (Bos taurus), sheep (Ovis aries) and goats (Capra hircus). In the bovine AQP10 gene, exon 1 and 5 had deletions resulting in a frame-shift or a premature termination, respectively, which were confirmed by the direct exon sequencing of the genomic DNA. In the RT-PCR experiments, the PCR primer sets for exon 1/2 and exon 4/5 failed to detect the bands for AQP10 mRNA in the duodenum and jejunum. Similar AQP10 gene mutations were also confirmed in the genomic DNA from sheep and goats. Although these animals were derived from porcine ancestors, the exons of the swine (Sus scrofa) AQP10 gene were complete without mutations. Therefore, AQP10 gene might have turned to a pseudogene around 65 million years before when cattle evolved from porcine ancestors. Conclusion AQP10 of ruminantia which regurgitate and rechew their food may have lost its role possibly due to the redundant expression of other aquaglyceroporins.

Aquaporin-11 (AQP11) Expression in the Mouse Brain

Aquaporin-11 (AQP11) is an intracellular aquaporin expressed in various tissues, including brain tissues in mammals. While AQP11-deficient mice have developed fatal polycystic kidneys at one month old, the role of AQP11 in the brain was not well appreciated. In this study, we examined the AQP11 expression in the mouse brain and the brain phenotype of AQP11-deficient mice. AQP11 messenger ribonucleic acid (mRNA) and protein were expressed in the brain, but much less than in the thymus and kidney. Immunostaining showed that AQP11 was localized at the epithelium of the choroid plexus and at the endothelium of the brain capillary, suggesting that AQP11 may be involved in water transport at the choroid plexus and blood-brain barrier (BBB) in the brain. The expression of AQP4, another brain AQP expressed at the BBB, was decreased by half in AQP11-deficient mice, thereby suggesting the presence of the interaction between AQP11 and AQP4. The brain of AQP11-deficient mice, however, did not show any morphological abnormalities and the function of the BBB was intact. Our findings provide a novel insight into a water transport mechanism mediated by AQPs in the brain, which may lead to a new therapy for brain edema.

Proteomic analysis of AQP11-null kidney: Proximal tubular type polycystic kidney disease

Autosomal Dominant Polycystic Kidney Disease (ADPKD) is caused by the mutation of polycystins (PC-1 or PC-2), in which cysts start from the collecting duct to extend to all nephron segments with eventual end stage renal failure. The cyst development is attenuated by a vasopressin V2 receptor antagonist tolvaptan which, however, will not affect proximal tubule cysts devoid of V2 receptor. Aquaporin-11 (AQP11) is expressed selectively in the proximal tubule of the kidney and AQP11-null kidneys have a disruptive PC-1 trafficking to the plasma membrane to develop polycystic kidneys. Here, we analyzed AQP11-null kidneys at the beginning of cyst formation by quantitative proteomic analysis using Tandem Mass Tag (TMT). Among ~ 1200 identified proteins, 124 proteins were differently expressed by > 1.5 or < 0.8 fold change. A pancreatic stone inhibitor or a growth factor, lithostathine-1 (Reg1) was most enhanced by 5 folds which was confirmed by western blot, while mitochondria-related proteins were downregulated. The identified proteins will be new target molecules for the treatment of proximal tubular cysts and helpful to explore the functional roles of AQP11 in the kidney.

Quantitative Analysis of Aquaporin Expression Levels during the Development and Maturation of the Inner Ear

Aquaporins (AQPs) are a family of small membrane proteins that transport water molecules across the plasma membrane along the osmotic gradient. Mammals express 13 subtypes of AQPs, including the recently reported “subcellular AQPs”, AQP11 and 12. Each organ expresses specific subsets of AQP subtypes, and in the inner ear, AQPs are essential for the establishment and maintenance of two distinct fluids, endolymph and perilymph. To evaluate the contribution of AQPs during the establishment of inner ear function, we used quantitative reverse transcription polymerase chain reaction to quantify the expression levels of all known AQPs during the entire development and maturation of the inner ear. Using systematic and longitudinal quantification, we found that AQP11 was majorly and constantly expressed in the inner ear, and that the expression levels of several AQPs follow characteristic longitudinal patterns: increasing (Aqp0, 1, and 9), decreasing (Aqp6, 8, and 12), and peak of expression on E18 (Aqp2, 5, and 7). In particular, the expression level of Aqp9 increased by 70-fold during P3–P21. We also performed in situ hybridization of Aqp11, and determined the unique localization of Aqp11 in the outer hair cells. Immunohistochemistry of AQP9 revealed its localization in the supporting cells inside the organ of Corti, and in the root cells. The emergence of AQP9 expression in these cells was during P3–P21, which was coincident with the marked increase of its expression level. Combining these quantification and localization data, we discuss the possible contributions of these AQPs to inner ear function.