14-3-3 proteins inactivate DAPK2 by promoting its dimerization and protecting key regulatory phosphosites.
14-3-3 proteins inactivate DAPK2 by promoting its dimerization and protecting key regulatory phosphosites.
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DOI：
10.1038/s42003-021-02518-y
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发表时间：
2021-08-19
影响因子：
5.9
通讯作者：
Obsil T
中科院分区：
文献类型：
--
作者：
Horvath M;Petrvalska O;Herman P;Obsilova V;Obsil T
死亡相关蛋白激酶2 （DAPK2）是cam调控的丝氨酸/苏氨酸蛋白激酶，参与细胞凋亡、自噬、粒细胞分化和运动调节，其活性受自抑制、自磷酸化、二聚化和与支架蛋白14-3-3的相互作用控制。然而，14-3-3介导的DAPK2调控的结构基础尚不清楚。本文结合多种生物物理技术，对全长人类DAPK2:14-3-3复合物进行了结构和生化表征。我们的x射线晶体分析结果显示，在DAPK2 C端Thr369磷酸化产生了一个高亲和力的典型模式III 14-3-3结合基元，二萜苷Fusicoccin a进一步增强了这一基元。此外，浓度依赖性的DAPK2二聚化被Ca2+/CaM结合破坏，并在溶液中被14-3-3结合稳定，从而保护了DAPK2抑制自磷酸化位点Ser318免受去磷酸化和阻止Ca2+/CaM结合。总的来说，我们的研究结果提供了14-3-3介导的DAPK2抑制的机制见解，并强调了DAPK2:14-3-3复合物作为抗炎治疗靶点的潜力。Horvath等人对全长人DAPK2-14-3-3复合物进行了结构和生化表征，研究了与DAPK2结合对其二聚化、Ser318去磷酸化激活以及Ca2+/钙调蛋白结合的影响。他们的研究结果为14-3-3介导的DAPK2抑制提供了机制见解，并强调了DAPK2:14-3-3复合物作为抗炎治疗靶点的潜力。
Death-associated protein kinase 2 (DAPK2) is a CaM-regulated Ser/Thr protein kinase, involved in apoptosis, autophagy, granulocyte differentiation and motility regulation, whose activity is controlled by autoinhibition, autophosphorylation, dimerization and interaction with scaffolding proteins 14-3-3. However, the structural basis of 14-3-3-mediated DAPK2 regulation remains unclear. Here, we structurally and biochemically characterize the full-length human DAPK2:14-3-3 complex by combining several biophysical techniques. The results from our X-ray crystallographic analysis revealed that Thr369 phosphorylation at the DAPK2 C terminus creates a high-affinity canonical mode III 14-3-3-binding motif, further enhanced by the diterpene glycoside Fusicoccin A. Moreover, concentration-dependent DAPK2 dimerization is disrupted by Ca2+/CaM binding and stabilized by 14-3-3 binding in solution, thereby protecting the DAPK2 inhibitory autophosphorylation site Ser318 against dephosphorylation and preventing Ca2+/CaM binding. Overall, our findings provide mechanistic insights into 14-3-3-mediated DAPK2 inhibition and highlight the potential of the DAPK2:14-3-3 complex as a target for anti‐inflammatory therapies. Horvath et al. structurally and biochemically characterize the full-length human DAPK2-14-3-3 complex to investigate the effects of binding to DAPK2 on its dimerization, activation by dephosphorylation of Ser318, and Ca2+/calmodulin binding. Their results provide mechanistic insights into 14- 3-3-mediated DAPK2 inhibition and highlight the potential of the DAPK2:14-3-3 complex as a target for anti-inflammatory therapies.