Subcellular location of serum- and glucocorticoid-induced kinase-1 in renal and mammary epithelial cells
Subcellular location of serum- and glucocorticoid-induced kinase-1 in renal and mammary epithelial cells
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DOI：
10.1152/ajpcell.00399.2006
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发表时间：
2007-05-01
影响因子：
5.5
通讯作者：
Fejes-Toth, Geza
中科院分区：
文献类型：
--
作者：
Cordas, Emily;Naray-Fejas-Toth, Aniko;Fejes-Toth, Geza
血清和糖皮质激素诱导的激酶-1（SGK 1）通过增加皮质集合管（CCD）细胞上皮Na+通道（ENaC）活性参与醛固酮诱导的Na+重吸收，但其确切作用机制尚不清楚。虽然在表达系统中已经描述了几个潜在的靶点，如Nedd 4 -2，但介导SGK 1生理作用的内源性底物仍有待鉴定。此外，SGK 1的亚细胞定位研究提供了有争议的结果。我们使用SGK 1-自身荧光蛋白（AFP）融合蛋白确定SGK 1的亚细胞定位。用编码SGK 1- AFP的构建体瞬时转染兔CCD（RCCT-28 A）细胞，并用各种亚细胞区室的标记物染色或共转染。在活细胞中，瞬时表达的SGK 1- AFP明显与线粒体标记物罗丹明123共定位。类似地，当使用逆转录病毒系统在RCCT-28 A细胞或乳腺上皮细胞系MCF 10A中稳定表达时，SGK 1- AFP与线粒体标记物MitoTracker共定位。为了确定SGK 1的哪个区域负责这种亚细胞定位，我们产生了稳定表达SGK 1突变体的RCCT-28 A细胞系。结果表明，SGK 1的NH 2端60个氨基酸的区域是其亚细胞定位所必需的和充分的。SGK 1定位于线粒体提高了SGK 1可能在调节能量代谢中发挥作用的可能性。
Serumand glucocorticoid- induced kinase- 1 ( SGK1) is involved in aldosterone- induced Na+ reabsorption by increasing epithelial Na+ channel ( ENaC) activity in cortical collecting duct ( CCD) cells, but its exact mechanisms of action are unknown. Although several potential targets such as Nedd4-2 have been described in expression systems, endogenous substrates mediating SGK1' s physiological effects remain to be identified. In addition, subcellular localization studies of SGK1 have provided controversial results. We determined the subcellular location of SGK1 using SGK1- autofluorescent protein ( AFP) fusion proteins. Rabbit CCD ( RCCT- 28A) cells were transiently transfected with a construct encoding for SGK1- AFP and were stained or cotransfected with markers for various subcellular compartments. In live cells, transiently expressed SGK1- AFP clearly colocalized with the mitochondrial marker rhodamine 123. Similarly, SGK1- AFP colocalized with the mitochondrial marker MitoTracker when stably expressed using a retroviral system in either RCCT- 28A cells or the mammary epithelial cell line MCF10A. To determine which region of SGK1 is responsible for this subcellular localization, we generated RCCT- 28A cell lines stably expressing SGK1 mutants. The results indicate that the NH2- terminal 60- amino acid region of SGK1 is necessary and sufficient for its subcellular localization. Localization of SGK1 to the mitochondria raises the possibility that SGK1 may play a role in regulating energy metabolism.