Uncoupling JAK2 V617F activation from cytokine-induced signalling by modulation of JH2 αC helix.
Uncoupling JAK2 V617F activation from cytokine-induced signalling by modulation of JH2 αC helix.
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DOI：
10.1042/bcj20160085
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发表时间：
2016-06-01
期刊：
影响因子：
--
通讯作者：
Constantinescu SN
中科院分区：
文献类型：
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作者：
Leroy E;Dusa A;Colau D;Motamedi A;Cahu X;Mouton C;Huang LJ;Shiau AK;Constantinescu SN
我们表明，假激酶 V617F 突变对 JAK2 的激活是由 JAK2 上的残基群介导的。假激酶螺旋 αC 上的电荷变化特异性抑制 V617F 诱导的激活。因此，变构调节可导致特异性 JAK2 V617F 抑制。血癌中常见的假激酶 (JH2) V617F 突变激活 JAK2 的机制仍不清楚。通过结构引导诱变以及转录和功能测定，我们鉴定了介导 V617F 诱导激活的 JH2 螺旋 αC、SH2-JH2 连接子和 JH1 激酶结构域的残基群。该电路通过改变 JH2 αC 溶剂暴露面的残基电荷来打破，预计 JH2 αC 会与 SH2-JH2 连接体和 JH1 相互作用。去除负电荷或添加正电荷的突变（例如 E596A/R）不会改变 JH2 V617F 折叠，如 JH2 V617F E596A 的晶体结构所示。相反，它们通过调节 SH2-JH2 接头的 C 端残基来防止激酶结构域激活。这些结果提出了选择性抑制 V617F JAK2 并保留野生型功能的策略。
We show that JAK2 activation by the pseudokinase V617F mutation is mediated by a community of residues across JAK2. Charge changes on pseudokinase helix αC specifically inhibits V617F-induced activation. Thus, allosteric modulation can lead to specific JAK2 V617F inhibition. The mechanisms by which JAK2 is activated by the prevalent pseudokinase (JH2) V617F mutation in blood cancers remain elusive. Via structure-guided mutagenesis and transcriptional and functional assays, we identify a community of residues from the JH2 helix αC, SH2-JH2 linker and JH1 kinase domain that mediate V617F-induced activation. This circuit is broken by altering the charge of residues along the solvent-exposed face of the JH2 αC, which is predicted to interact with the SH2-JH2 linker and JH1. Mutations that remove negative charges or add positive charges, such as E596A/R, do not alter the JH2 V617F fold, as shown by the crystal structure of JH2 V617F E596A. Instead, they prevent kinase domain activation via modulation of the C-terminal residues of the SH2-JH2 linker. These results suggest strategies for selective V617F JAK2 inhibition, with preservation of wild-type function.