Structural and biochemical characterization of the KLHL3-WNK kinase interaction important in blood pressure regulation.
Structural and biochemical characterization of the KLHL3-WNK kinase interaction important in blood pressure regulation.
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DOI：
10.1042/bj20140153
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发表时间：
2014-06-01
期刊：
影响因子：
--
通讯作者：
Kurz T
中科院分区：
文献类型：
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作者：
Schumacher FR;Sorrell FJ;Alessi DR;Bullock AN;Kurz T
WNK1 [不含赖氨酸 (K)] 和 WNK4 通过控制肾脏中离子协同转运蛋白的活性来调节血压。突破性的工作表明，WNK 亚型的泛素化及其水平是由 Cullin-RING E3 泛素连接酶复合物 (CRL3KLHL3) 控制的，该复合物利用 CUL3 (Cullin3) 及其底物接头 KLHL3（Kelch 样蛋白 3）。 CUL3 或 KLHL3 的功能丧失突变通过稳定 WNK 亚型导致遗传性高血压疾病戈登氏综合征。 KLHL3 与位于 WNK 亚型 C 端非催化结构域内的高度保守的降解决定子基序结合。这种相互作用对于 CRL3KLHL3 的泛素化至关重要，WNK4 和 KLHL3 的致病突变通过破坏这种相互作用来对血压产生影响。在本研究中，我们报告了与 WNK4 降解决定子基序复合的 KLHL3 Kelch 结构域的晶体结构。这揭示了 Kelch 结构域 β-螺旋桨表面的保守残基与 WNK4 降解决定子基序之间复杂的相互作用网络。重要的是，许多致病突变通过破坏关键的界面接触来抑制结合。我们还展示了与 KLHL2 复合的 WNK4 降解决定子基序的结构，据报道 KLHL2 也能结合 WNK4。这证实了 KLHL2 与 WNK 激酶的相互作用方式与 KLHL3 类似，但与另一种 KLHL 蛋白 KEAP1（Kelch 样烯酰基辅酶 A 水合酶相关蛋白 1）与其底物 NRF2（核因子 - 红细胞 2 相关因子 2）的结合方式截然不同。本研究进一步深入了解了 Kelch 样衔接蛋白如何识别其底物，并为 WNK4 和 KLHL3 突变如何导致高血压提供了结构基础。 WNK 激酶调节哺乳动物血压。细胞中 WNK 蛋白的水平由 KLHL3–CUL3 泛素连接酶调节。我们定义了 KLHL3 和 WNK 之间的相互作用，识别了 WNK 降解决定子，并展示了 KLHL3-WNK 降解决定子复合物的晶体结构。
WNK1 [with no lysine (K)] and WNK4 regulate blood pressure by controlling the activity of ion co-transporters in the kidney. Groundbreaking work has revealed that the ubiquitylation and hence levels of WNK isoforms are controlled by a Cullin-RING E3 ubiquitin ligase complex (CRL3KLHL3) that utilizes CUL3 (Cullin3) and its substrate adaptor, KLHL3 (Kelch-like protein 3). Loss-of-function mutations in either CUL3 or KLHL3 cause the hereditary high blood pressure disease Gordon's syndrome by stabilizing WNK isoforms. KLHL3 binds to a highly conserved degron motif located within the C-terminal non-catalytic domain of WNK isoforms. This interaction is essential for ubiquitylation by CRL3KLHL3 and disease-causing mutations in WNK4 and KLHL3 exert their effects on blood pressure by disrupting this interaction. In the present study, we report on the crystal structure of the KLHL3 Kelch domain in complex with the WNK4 degron motif. This reveals an intricate web of interactions between conserved residues on the surface of the Kelch domain β-propeller and the WNK4 degron motif. Importantly, many of the disease-causing mutations inhibit binding by disrupting critical interface contacts. We also present the structure of the WNK4 degron motif in complex with KLHL2 that has also been reported to bind WNK4. This confirms that KLHL2 interacts with WNK kinases in a similar manner to KLHL3, but strikingly different to how another KLHL protein, KEAP1 (Kelch-like enoyl-CoA hydratase-associated protein 1), binds to its substrate NRF2 (nuclear factor-erythroid 2-related factor 2). The present study provides further insights into how Kelch-like adaptor proteins recognize their substrates and provides a structural basis for how mutations in WNK4 and KLHL3 lead to hypertension. WNK kinases regulate mammalian blood pressure. The level of WNK protein in a cell is regulated by the KLHL3–CUL3 ubiquitin ligase. We define the interaction between KLHL3 and WNK, identifying the WNK degron, and present the crystal structure of the KLHL3–WNK degron complex.