Institute for Integrated Cell-Material Sciences (WPI-iCeMS), Kyoto University, Yoshida-Honmachi, Sakyoku, Kyoto 606-8501, Japan; Center for Integrated Biosystems, Institute for Biomedical Sciences, Academia Sinica, Taipei, Taiwan.
Institute for Integrated Cell-Material Sciences (WPI-iCeMS), Kyoto University, Yoshida-Honmachi, Sakyoku, Kyoto 606-8501, Japan; Graduate School of Biostudies, Kyoto University, Konoe-cho, Yoshida, Sakyoku, Kyoto 606-8501, Japan.
Mol Cell. 2021 Apr 1;81(7):1397-1410.e9. doi: 10.1016/j.molcel.2021.02.025. Epub 2021 Mar 15.
Phospholipid scrambling in dying cells promotes phosphatidylserine exposure, a critical process for efferocytosis. We previously identified the Xkr family protein Xkr4 as a phospholipid-scrambling protein, but its activation mechanisms remain unknown. Here we show that Xkr4 is activated in two steps: dimer formation by caspase-mediated cleavage and structural change caused by activating factors. To identify the factors, we developed a new screening system, "revival screening," using a CRISPR sgRNA library. Applying this system, we identified the nuclear protein XRCC4 as the single candidate for the Xkr4 activator. Upon apoptotic stimuli, XRCC4, contained in the DNA repair complex, is cleaved by caspases, and its C-terminal fragment with an intrinsically disordered region is released into the cytoplasm. Protein interaction screening showed that the fragment interacts directly with the Xkr4 dimer to activate it. This study demonstrates that caspase-mediated cleavage releases a nuclear protein fragment for direct regulation of lipid dynamics on the plasma membrane.
在死亡细胞中，磷脂重排促进磷脂酰丝氨酸暴露，这是细胞吞噬作用的关键过程。我们之前发现 Xkr 家族蛋白 Xkr4 是一种磷脂重排蛋白，但它的激活机制尚不清楚。在这里，我们表明 Xkr4 的激活分为两步：半胱天冬酶介导的切割形成二聚体和激活因子引起的结构变化。为了鉴定这些因子，我们开发了一种新的筛选系统，“复苏筛选”，使用了 CRISPR sgRNA 文库。应用该系统，我们鉴定了核蛋白 XRCC4 是 Xkr4 激活剂的唯一候选物。在凋亡刺激下，包含在 DNA 修复复合物中的 XRCC4 被半胱天冬酶切割，其带有无规则结构域的 C 末端片段被释放到细胞质中。蛋白质相互作用筛选表明，该片段直接与 Xkr4 二聚体相互作用，从而激活 Xkr4。这项研究表明，半胱天冬酶介导的切割释放了核蛋白片段，可直接调节质膜上的脂质动态。
