Aquaporin-2 regulates cell volume recovery via tropomyosin.
Cell volume regulation is particularly important for kidney collecting duct cells. These cells are the site of water reabsorption regulated by vasopressin and aquaporin-2 (AQP2) trafficking to the apical membrane, and subject to changes in osmolality. Here, we examined the role of AQP2 in regulatory volume decrease (RVD), which is a cellular defensive process against hypotonic stress. Stable expression of AQP2 increases RVD in MDCK cells and its phosphorylation levels decrease during the RVD process. We then examined the involvement of AQP2 phosphorylation at serine 256 and serine 261 in RVD using cells stably expressing the phosphorylation mutants. Both S256A- and S256D-AQP2 decrease RVD compared to wild type (WT)-AQP2 although only S256A mutation decreases the initial osmotic swelling, indicating that AQP2-enhanced RVD is independent of osmotic swelling induced by the water permeability of AQP2. S261A and S261D mutations do not induce changes compared with WT-AQP2. These findings indicate that switching between phosphorylation and dephosphorylation at S256 is important for RVD. We previously reported that AQP2 interacts with tropomyosin 5b (TM5b), which regulates actin stability. AQP2 interactions with TM5b are rapidly increased by hypotonicity and then decreased, which are consistent with AQP2 phosphorylation levels. Knockdown and overexpression of TM5b show its essential role in WT-AQP2-enhanced RVD. RVD in S256A- and S256D-AQP2-expressing cells is not changed by TM5b knockdown or overexpression. The present study shows that AQP2 regulates RVD via TM5b and switching between phosphorylation and dephosphorylation at S256 in AQP2 is critical for this process.
Li
Yu-Hua
YH
Department of Pharmacology, School of Basic Medical Sciences, Peking University, Beijing 100191, China.
Eto
Kayoko
K
Horikawa
Saburo
S
Uchida
Shinichi
S
Sasaki
Sei
S
Li
Xue-Jun
XJ
Noda
Yumi
Y
D000818:Animals; D051399:Aquaporin 2 / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D002462:Cell Membrane; D002463:Cell Membrane Permeability; D048429:Cell Size; D003001:Cloning, Molecular; D004285:Dogs; D007685:Kidney Tubules, Collecting / Q000166:cytology / Q000378:metabolism*; D050505:Mutant Proteins / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation; D021381:Protein Transport; D034741:RNA, Small Interfering / Q000235:genetics; D019076:Transgenes / Q000235:genetics; D014335:Tropomyosin / Q000235:genetics / Q000378:metabolism
D051399:Aquaporin 2; D050505:Mutant Proteins; D034741:RNA, Small Interfering; D014335:Tropomyosin