Osmosensing by WNK Kinases.
Osmosensing by WNK Kinases.
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DOI：
10.1091/mbc.e20-01-0089
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发表时间：
2021-08-19
影响因子：
3.3
通讯作者：
Goldsmith EJ
中科院分区：
文献类型：
--
作者：
Akella R;Humphreys JM;Sekulski K;He H;Durbacz M;Chakravarthy S;Liwocha J;Mohammed ZJ;Brautigam CA;Goldsmith EJ
在无赖氨酸（K）的情况下，WNK激酶调节由渗透胁迫和氯控制的电中性共转运蛋白。我们先前表明，WNK 1的自磷酸化被氯化物抑制，提高了WNK被渗透胁迫激活的可能性。在这里，我们证明了未磷酸化的WNK亚型3和1 autophosphorylate响应于渗透压在体外，应用拥挤剂聚乙二醇（PEG）400或渗透剂乙二醇（EG），这种激活是反对氯化物。在存在和不存在PEG 400的情况下WNK 3的小角X射线散射、EG中的静态光散射和WNK 1的晶体学用于理解该机制。WNK 3和WNK 1中的渗透传感似乎是通过无活性的、未磷酸化的氯离子结合二聚体和自磷酸化单体之间的构象平衡发生的。WNK 1的非活性激酶结构域的改进结构，以及与WNK 1的单磷酸化形式的结构的比较表明，大空腔、更大的水合作用和特定的结合水可能参与了WNK 1的生物传感机制。我们先前的工作表明，渗透调节剂对磷酸化WNK 1的结构有影响，提示WNK中的渗透调节有多个阶段。
With No Lysine (K) WNK kinases regulate electro-neutral cotransporters that are controlled by osmotic stress and chloride. We showed previously that autophosphorylation of WNK1 is inhibited by chloride, raising the possibility that WNKs are activated by osmotic stress. Here we demonstrate that unphosphorylated WNK isoforms 3 and 1 autophosphorylate in response to osmotic pressure in vitro, applied with the crowding agent polyethylene glycol (PEG)400 or osmolyte ethylene glycol (EG), and that this activation is opposed by chloride. Small angle x-ray scattering of WNK3 in the presence and absence of PEG400, static light scattering in EG, and crystallography of WNK1 were used to understand the mechanism. Osmosensing in WNK3 and WNK1 appears to occur through a conformational equilibrium between an inactive, unphosphorylated, chloride-binding dimer and an autophosphorylation-competent monomer. An improved structure of the inactive kinase domain of WNK1, and a comparison with the structure of a monophosphorylated form of WNK1, suggests that large cavities, greater hydration, and specific bound water may participate in the osmosensing mechanism. Our prior work showed that osmolytes have effects on the structure of phosphorylated WNK1, suggestive of multiple stages of osmotic regulation in WNKs.