Regulation of the water channel aquaporin-2 by cullin E3 ubiquitin ligases.
Regulation of the water channel aquaporin-2 by cullin E3 ubiquitin ligases.
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cullin E3 泛素连接酶对水通道 aquaporin-2 的调节。
DOI：
10.1152/ajprenal.00049.2024
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发表时间：
2024
期刊：
影响因子：
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通讯作者：
Fenton,RobertA
中科院分区：
文献类型：
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作者：
Murali,SathishK;McCormick,JamesA;Fenton,RobertA
水通道蛋白2（AQP 2）是肾脏集合管中由血管加压素（VP）调节的水通道。AQP 2的磷酸化和泛素化在控制细胞内AQP 2的丰度及其在细胞膜上的积累中起重要作用。Cullin-RING泛素连接酶（CRL）是参与其靶蛋白的泛素化和降解的多亚基E3连接酶，其中8种在集合管中表达。在这里，我们使用一个已建立的集合管细胞模型（mpkCCD 14细胞），研究cullin在调节AQP 2中的作用。蛋白质印迹鉴定了mpkCCD 14细胞中的Cul-1至Cul-5。与溶剂相比，用泛cullin抑制剂（MLN 4924）处理细胞4 h可降低AQP 2丰度，阻止VP诱导的AQP 2 Ser 261磷酸化减少，并减弱VP诱导的AQP 2质膜蓄积。与对照组相比，MLN 4924给药后AQP 2泛素化水平显著升高，尽管VP给药，其仍保持较高水平。Cullin抑制增加ERK 1/2活性（一种调节AQP 2 Ser 261磷酸化的激酶），MLN 4924处理期间不存在VP诱导的ERK 1/2磷酸化降低。此外，MLN 4924处理期间Ser 261磷酸化增加和AQP 2丰度降低在ERK 1/2抑制期间减弱。MLN 4924通过钙释放激活的钙通道增加细胞内钙水平，抑制该通道可消除MLN 4924对Ser 261磷酸化和AQP 2丰度的影响。总之，CRL在介导VP增加AQP 2质膜积累和AQP 2丰度的一些作用中起着至关重要的作用。调节cullin活性是否有助于体内水分平衡还需要进一步的研究。新&值得注意的是水通道蛋白2（AQP 2）对体内水分平衡至关重要，并受抗利尿激素加压素调节。翻译后修饰泛素化是AQP 2丰度和质膜定位的关键调节因子。在这里，我们证明了cullin-RING E3连接酶在介导加压素增加AQP 2丰度和质膜积累的一些作用中起着至关重要的作用。结果表明，操纵cullin活性可能是改变肾脏水处理的一种新策略。
Aquaporin 2 (AQP2) is a vasopressin (VP)-regulated water channel in the renal collecting duct. Phosphorylation and ubiquitylation of AQP2 play an essential role in controlling the cellular abundance of AQP2 and its accumulation on the plasma membrane in response to VP. Cullin-RING ubiquitin ligases (CRLs) are multisubunit E3 ligases involved in ubiquitylation and degradation of their target proteins, eight of which are expressed in the collecting duct. Here, we used an established cell model of the collecting duct (mpkCCD14 cells) to study the role of cullins in modulating AQP2. Western blotting identified Cul-1 to Cul-5 in mpkCCD14 cells. Treatment of cells for 4 h with a pan-cullin inhibitor (MLN4924) decreased AQP2 abundance, prevented a VP-induced reduction in AQP2 Ser261phosphorylation, and attenuated VP-induced plasma membrane accumulation of AQP2 relative to the vehicle. AQP2 ubiquitylation levels were significantly higher after MLN4924 treatment compared with controls, and they remained higher despite VP treatment. Cullin inhibition increased ERK1/2 activity, a kinase that regulates AQP2 Ser261phosphorylation, and VP-induced reductions in ERK1/2 phosphorylation were absent during MLN4924 treatment. Furthermore, the greater Ser261phosphorylation and reduction in AQP2 abundance during MLN4924 treatment were attenuated during ERK1/2 inhibition. MLN4924 increased intracellular calcium levels via calcium release-activated calcium channels, inhibition of which abolished MLN4924 effects on Ser261phosphorylation and AQP2 abundance. In conclusion, CRLs play a vital role in mediating some of the effects of VP to increase AQP2 plasma membrane accumulation and AQP2 abundance. Whether modulation of cullin activity can contribute to body water homeostasis requires further studies.NEW & NOTEWORTHYAquaporin 2 (AQP2) is essential for body water homeostasis and is regulated by the antidiuretic hormone vasopressin. The posttranslational modification ubiquitylation is a key regulator of AQP2 abundance and plasma membrane localization. Here we demonstrate that cullin-RING E3 ligases play a vital role in mediating some of the effects of vasopressin to increase AQP2 abundance and plasma membrane accumulation. The results suggest that manipulating cullin activity could be a novel strategy to alter kidney water handling.