Ubiquitination of NKCC2 by the cullin-RING E3 ubiquitin ligase family in the thick ascending limb of the loop of Henle.
Ubiquitination of NKCC2 by the cullin-RING E3 ubiquitin ligase family in the thick ascending limb of the loop of Henle.
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DOI：
10.1152/ajprenal.00079.2022
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发表时间：
2023-03-01
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--
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在亨利氏袢（TAL）的粗升支中的Na+/K+/2Cl−协同转运蛋白（NKCC 2）介导NaCl的重吸收。cGMP是一氧化氮和心房利钠肽的第二信使，通过刺激NKCC 2泛素化和降低表面NKCC 2水平来抑制NKCC 2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。Cullin是分子支架蛋白，其募集多个亚基以形成CRL复合物。我们假设CRL复合物介导TAL中NKCC 2泛素化的cGMP依赖性增加。在大鼠TAL中，Cullin-1、cullin-2、cullin-3、cullin-4A和cullin-5在蛋白水平上表达，而cullin家族的其他成员在mRNA水平上表达。CRL复合物活性由神经元前体细胞表达的发育下调蛋白8（Nedd 8）调节为cullin，这一过程称为neddylation。抑制cullin neddylation减弱了泛素化NKCC 2的cGMP依赖性增加，同时cullin-1的表达增加了3倍，但这种作用在其他cullin中没有观察到。CRL复合物的活性也受cullin相关的Nedd 8-解离1（CAND 1）的调节。CAND 1与cullin结合，促进底物识别蛋白的交换，以靶向不同的蛋白质进行泛素化。CAND 1抑制加剧了NKCC 2泛素化的cGMP依赖性增加，并降低了表面NKCC 2表达。最后，cGMP增加cullin的neddylation。我们的结论是cGMP依赖性增加NKCC 2泛素化是由CRL复合物介导的。据我们所知，这是CRL复合物介导天然TAL中NKCC 2泛素化的第一个证据。Na+/K+/2Cl−协同转运蛋白（NKCC 2）通过粗大的上升支重吸收NaCl。一氧化氮和心房利钠肽通过增加第二信使cGMP减少粗大的上行肢体对NaCl的重吸收。目前的研究结果表明，cGMP通过cullin-RING连接酶复合物增加NKCC 2泛素化，并部分调节表面NKCC 2水平。鉴定调节NKCC 2表达和活性的E3泛素连接酶可能为开发特异性袢利尿剂提供新的靶点。
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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影响因子：
64.8
作者：
Boyden, Lynn M.;Choi, Murim;Choate, Keith A.;Nelson-Williams, Carol J.;Farhi, Anita;Toka, Hakan R.;Tikhonova, Irina R.;Bjornson, Robert;Mane, Shrikant M.;Colussi, Giacomo;Lebel, Marcel;Gordon, Richard D.;Semmekrot, Ben A.;Poujol, Alain;Valimaki, Matti J.;De Ferrari, Maria E.;Sanjad, Sami A.;Gutkin, Michael;Karet, Fiona E.;Tucci, Joseph R.;Stockigt, Jim R.;Keppler-Noreuil, Kim M.;Porter, Craig C.;Anand, Sudhir K.;Whiteford, Margo L.;Davis, Ira D.;Dewar, Stephanie B.;Bettinelli, Alberto;Fadrowski, Jeffrey J.;Belsha, Craig W.;Hunley, Tracy E.;Nelson, Raoul D.;Trachtman, Howard;Cole, Trevor R. P.;Pinsk, Maury;Bockenhauer, Detlef;Shenoy, Mohan;Vaidyanathan, Priya;Foreman, John W.;Rasoulpour, Majid;Thameem, Farook;Al-Shahrouri, Hania Z.;Radhakrishnan, Jai;Gharavi, Ali G.;Goilav, Beatrice;Lifton, Richard P.
通讯作者：
Lifton, Richard P.
影响因子：
6.8
作者：
Duda DM;Scott DC;Calabrese MF;Zimmerman ES;Zheng N;Schulman BA
通讯作者：
Schulman BA
影响因子：
30.8
作者：
HANSSON, JH;NELSONWILLIAMS, C;LIFTON, RP
通讯作者：
LIFTON, RP
影响因子：
8.3
作者：
Hoagland, KM;Flasch, AK;Roman, RJ
通讯作者：
Roman, RJ
DOI：
10.1016/j.bbrc.2016.10.127
发表时间：
2016-11
影响因子：
3.1
作者：
Ishizawa, Kenichi;Xu, Ning;Loffing, Johannes;Lifton, Richard P.;Fujita, Toshiro;Uchida, Shunya;Shibata, Shigeru
通讯作者：
Shibata, Shigeru