Glycogen Synthase Kinase-3 Modulates Hyperosmotic-Induced Urea Transporter A1 Relocation in the Inner Medullary Collecting Duct Cells
Glycogen Synthase Kinase-3 Modulates Hyperosmotic-Induced Urea Transporter A1 Relocation in the Inner Medullary Collecting Duct Cells
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糖原合酶激酶 3 调节高渗诱导的尿素转运蛋白 A1 在内髓集合管细胞中的重新定位
作者：
Li Yong-xia;Huang Yun;Liu Song;Mao Yan;Yuan Cheng-yan;Yang Xiao;Yao Li-jun
目的：糖原合成酶激酶3（GSK 3）通过介导加压素诱导的水通道蛋白2表达和水渗透性来调节尿液浓度，尽管GSK 3是否也介导尿素转运蛋白A1（UT-A1）的积累尚不清楚。本研究的目的是研究GSK 3对UT-A1分布的影响。方法：将UT-A1-GFP转染小鼠内髓集合管3细胞。将稳定转染的细胞在高渗条件下培养，用GSK 3抑制剂氯化锂、GSK 3激活剂、溶酶体或蛋白酶体抑制剂处理。使用蛋白质印迹分析UT-A1、GSK 3和磷酸化-GSK 3的表达水平。共聚焦免疫荧光显微镜观察UT-A1与高尔基体的相互作用。UT-A1运输使用表面膜的生物素化进行检查。结果：在高渗NaCl和NaCl+尿素刺激下，UT-A1从高尔基体解离并移位至质膜。这种运动伴随着GSK 3磷酸化的增加及其在细胞膜上的定位。此外，这些结果通过用GSK 3抑制剂处理细胞而重复，并且相反，被GSK 3激活剂部分逆转。用溶酶体或蛋白酶体抑制剂处理细胞未能减弱高渗刺激的影响，表明高尔基体UT-A1的丢失不是由于降解。结论：我们的研究结果表明，GSK 3可能部分调节高渗诱导的细胞内UT-A1的再分布及其在质膜上的积累，这可能构成GSK 3调节尿浓度的另一种机制。
Aim: Glycogen synthase kinase 3 (GSK3) regulates urine concentration by mediating the vasopressin-induced aquaporin 2 expression and water permeability, although it is unknown whether GSK3 also mediates the accumulation of the urea transporter A1 (UT-A1). The aim of this study is to investigate the effect of GSK3 on UT-A1 distribution. Methods: Mouse inner medullary collecting duct 3 cells were transfected with UT-A1-GFP construct. The stable transfected cells were cultured under hypertonic conditions, treated with GSK3 inhibitor lithium chloride, GSK3 activator, lysosome or proteasome inhibitor. The expression levels of UT-A1, GSK3, and phospho-GSK3 were analyzed using western blot. The interaction between UT-A1 and the Golgi apparatus was examined using confocal immunofluorescence microscope. The UT-A1 trafficking was examined using the biotinylation of surface membranes. Results: UT-A1 dissociated away from the Golgi apparatus and translocated to the plasma membrane under hypertonic-NaCl and NaCl plus urea stimulation. This movement was accompanied by the increased phosphorylation of GSK3 and its localization on the cellular membrane. Moreover, these results were duplicated by treating the cells with the GSK3 inhibitor, and by contrast, were partially reversed by the GSK3 activator. Treating cells with a lysosome or proteasome inhibitor failed to attenuate the effects of hypertonic stimulus, indicating that the loss of UT-A1 from the Golgi was not due to degradation. Conclusion: Our results suggest that GSK3 may in part modulate the hypertonic-induced intracellular UT-A1 redistribution and its accumulation on the plasma membrane, which may constitute another mechanism by which GSK3 modulates urine concentration.
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DOI：
10.1152/ajprenal.00664.2011
发表时间：
2012-05
期刊：
American journal of physiology. Renal physiology
影响因子：
--
作者：
J. Klein;Christopher F. Martin;Kimilia J Kent;J. Sands
通讯作者：
J. Klein;Christopher F. Martin;Kimilia J Kent;J. Sands
DOI：
10.1083/jcb.131.6.1715
发表时间：
1995-12
期刊：
The Journal of cell biology
影响因子：
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作者：
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通讯作者：
Warren G
影响因子：
3.3
作者：
Davanger, S;Bock, JB;Scheller, RH
通讯作者：
Scheller, RH
DOI：
10.1681/asn.2009060672
发表时间：
2010-03
期刊：
Journal of the American Society of Nephrology : JASN
影响因子：
--
作者：
Reena Rao;Satish Patel;C. Hao;J. Woodgett;Raymond C. Harris
通讯作者：
Reena Rao;Satish Patel;C. Hao;J. Woodgett;Raymond C. Harris
DOI：
10.1152/ajprenal.1998.274.6.f1167
发表时间：
1998-06
期刊：
American journal of physiology. Renal physiology
影响因子：
--
作者：
B. Santos;A. Chevaile;M. Hébert;J. Zagajeski;S. Gullans
通讯作者：
B. Santos;A. Chevaile;M. Hébert;J. Zagajeski;S. Gullans