Autophosphorylation transforms DNA-PK from protecting to processing DNA ends.
Autophosphorylation transforms DNA-PK from protecting to processing DNA ends.
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自身磷酸化使DNA-PK从保护DNA末端转变为处理DNA末端。
DOI：
10.1016/j.molcel.2021.11.025
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发表时间：
2022-01-06
期刊：
影响因子：
16
通讯作者：
Gellert M
中科院分区：
文献类型：
--
作者：
Liu L;Chen X;Li J;Wang H;Buehl CJ;Goff NJ;Meek K;Yang W;Gellert M
DNA依赖性蛋白激酶（DNA-PK）最初保护断裂的DNA末端，但随后在非同源末端连接（NHEJ）期间促进其加工。在通过NHEJ连接之前，在V（D）J重组期间产生的DNA发夹末端必须被Artemis核酸酶连同自磷酸化的DNA-PK一起打开。DNA-PK在磷酸化之前和之后与DNA结合的结构，以及与Artemis和DNA发夹的复合物，揭示了一个重要的功能开关。当在其保护模式下与开放DNA末端结合时，DNA-PK被抑制用于所谓的ABCDE簇的顺式自磷酸化，但被激活用于其他靶标的磷酸化。相反，DNA发夹末端促进顺式自磷酸化。ABCDE中四个Thr残基的磷酸化导致DNA-PK的总体结构重排，拓宽了Artemis募集和发夹切割的DNA结合沟。同时，Artemis将DNA-PK锁定在激酶失活状态。激酶活性和DNA-PK的自磷酸化由不同的DNA末端调节，前馈以协调NHEJ事件。Liu等人发现，与平端DNA结合的DNA-PK可以顺式自磷酸化（约33%）或反式磷酸化其他靶标（67%），同时保护DNA末端。DNA发夹使平衡向自磷酸化方向移动，导致DNA-PK的急剧构象变化，暴露DNA末端，Artemis募集和激酶自抑制。
The DNA-dependent protein kinase (DNA-PK) initially protects broken DNA ends, but then promotes their processing during non-homologous end-joining (NHEJ). Before ligation by NHEJ, DNA hairpin ends generated during V(D)J recombination must be opened by the Artemis nuclease together with autophosphorylated DNA-PK. Structures of DNA-PK bound to DNA before and after phosphorylation, and in complex with Artemis and a DNA hairpin, reveal an essential functional switch. When bound to open DNA ends in its protection mode, DNA-PK is inhibited for cis-autophosphorylation of the so-called ABCDE cluster but activated for phosphorylation of other targets. In contrast, DNA hairpin ends promote cis-autophosphorylation. Phosphorylation of four Thr residues in ABCDE leads to gross structural rearrangement of DNA-PK, widening the DNA-binding groove for Artemis recruitment and hairpin cleavage. Meanwhile, Artemis locks DNA-PK into the kinase-inactive state. Kinase activity and autophosphorylation of DNA-PK are regulated by different DNA ends, feeding forward to coordinate NHEJ events. Liu et al. find that DNA-PK bound to a blunt-end DNA can autophosphorylate in cis (~33%) or phosphorylate other targets in trans (67%) while protecting the DNA end. A DNA hairpin shifts the equilibrium toward autophosphorylation, leading to dramatic conformational changes of DNA-PK, exposing DNA-end, Artemis recruitment, and kinase self-inhibition.
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