cullin-RING E3泛素连接酶家族在Henle环厚升肢中的NKCC2泛素化。

IF 3.7 2区 医学 Q1 PHYSIOLOGY American Journal of Physiology-renal Physiology Pub Date : 2023-03-01 DOI:10.1152/ajprenal.00079.2022
Gustavo R Ares
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引用次数: 0

摘要

Henle环(TAL)厚升肢的Na+/K+/2Cl-共转运体(NKCC2)介导NaCl重吸收。cGMP是一氧化氮和房利钠肽的第二信使,通过刺激NKCC2泛素化和降低表面NKCC2水平来抑制NKCC2活性。在E3泛素连接酶家族中,cullin-RING E3泛素连接酶(CRL)家族是最大的。Cullins是招募多个亚基形成CRL复合物的分子支架蛋白。我们假设CRL复合物介导cgmp依赖性的NKCC2泛素化增加。cullin- 1、cullin-2、cullin-3、cullin- 4a和cullin-5在大鼠tal中以蛋白水平表达,而cullin家族的其他成员以mRNA水平表达。CRL复合物的活性受神经元前体细胞表达的发育下调蛋白8 (Nedd8)对cullins的调节,这一过程称为类化修饰。抑制cullin泛素化抑制了cgmp依赖性的泛素化NKCC2的增加,同时使cullin-1的表达增加了三倍,但在其他cullin中没有看到这种影响。CRL复合物的活性也受cullin-associated Nedd8-dissociated 1 (CAND1)的调控。CAND1结合cullins并促进底物识别蛋白的交换,以靶蛋白泛素化。CAND1抑制加剧了cgmp依赖性NKCC2泛素化的增加和表面NKCC2表达的降低。最后,cGMP增加了cullins的类化修饰。我们得出结论,cgmp依赖性的NKCC2泛素化增加是由CRL复合物介导的。据我们所知,这是CRL复合物介导天然tal中NKCC2泛素化的第一个证据。Na+/K+/2Cl-共转运体(NKCC2)通过厚升肢重吸收NaCl。一氧化氮和房利钠肽通过增加第二信使cGMP来降低粗升肢的NaCl重吸收。目前的研究结果表明,cGMP通过cullin-RING连接酶复合物增加NKCC2的泛素化,并在一定程度上调节表面NKCC2的水平。确定调节NKCC2表达和活性的E3泛素连接酶可能为开发特异性环利尿剂提供新的靶点。
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Ubiquitination of NKCC2 by the cullin-RING E3 ubiquitin ligase family in the thick ascending limb of the loop of Henle.

The Na+/K+/2Cl- cotransporter (NKCC2) in the thick ascending limb of the loop of Henle (TAL) mediates NaCl reabsorption. cGMP, the second messenger of nitric oxide and atrial natriuretic peptide, inhibits NKCC2 activity by stimulating NKCC2 ubiquitination and decreasing surface NKCC2 levels. Among the E3 ubiquitin ligase families, the cullin-RING E3 ubiquitin ligase (CRL) family is the largest. Cullins are molecular scaffold proteins that recruit multiple subunits to form the CRL complex. We hypothesized that a CRL complex mediates the cGMP-dependent increase in NKCC2 ubiquitination in TALs. Cullin-1, cullin-2, cullin-3, cullin-4A, and cullin-5 were expressed at the protein level, whereas the other members of the cullin family were expressed at the mRNA level, in rat TALs. CRL complex activity is regulated by neuronal precursor cell-expressed developmentally downregulated protein 8 (Nedd8) to cullins, a process called neddylation. Inhibition of cullin neddylation blunted the cGMP-dependent increase in ubiquitinated NKCC2 while increasing the expression of cullin-1 by threefold, but this effect was not seen with other cullins. CRL complex activity is also regulated by cullin-associated Nedd8-dissociated 1 (CAND1). CAND1 binds to cullins and promotes the exchange of substrate-recognition proteins to target different proteins for ubiquitination. CAND1 inhibition exacerbated the cGMP-dependent increase in NKCC2 ubiquitination and decreased surface NKCC2 expression. Finally, cGMP increased neddylation of cullins. We conclude that the cGMP-dependent increase in NKCC2 ubiquitination is mediated by a CRL complex. To the best of our knowledge, this is the first evidence that a CRL complex mediates NKCC2 ubiquitination in native TALs.NEW & NOTEWORTHY The Na+/K+/2Cl- cotransporter (NKCC2) reabsorbs NaCl by the thick ascending limb. Nitric oxide and atrial natriuretic peptide decrease NaCl reabsorption in thick ascending limbs by increasing the second messenger cGMP. The present findings indicate that cGMP increases NKCC2 ubiquitination via a cullin-RING ligase complex and regulates in part surface NKCC2 levels. Identifying the E3 ubiquitin ligases that regulate NKCC2 expression and activity may provide new targets for the development of specific loop diuretics.

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来源期刊
CiteScore
8.40
自引率
7.10%
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154
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2-4 weeks
期刊介绍: The American Journal of Physiology - Renal Physiology publishes original manuscripts on timely topics in both basic science and clinical research. Published articles address a broad range of subjects relating to the kidney and urinary tract, and may involve human or animal models, individual cell types, and isolated membrane systems. Also covered are the pathophysiological basis of renal disease processes, regulation of body fluids, and clinical research that provides mechanistic insights. Studies of renal function may be conducted using a wide range of approaches, such as biochemistry, immunology, genetics, mathematical modeling, molecular biology, as well as physiological and clinical methodologies.
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