Mechanism and cellular function of direct membrane binding by the ESCRT and ERES-associated Ca2+-sensor ALG-2.
Mechanism and cellular function of direct membrane binding by the ESCRT and ERES-associated Ca2+-sensor ALG-2.
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ESCRT 和 ERES 相关 Ca2 传感器 ALG-2 直接膜结合的机制和细胞功能。
DOI：
10.1101/2023.10.17.562764
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发表时间：
2023
期刊：
影响因子：
--
通讯作者：
Hurley,JamesH
中科院分区：
文献类型：
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作者：
Shukla,Sankalp;Chen,Wei;Rao,Shanlin;Yang,Serim;Ou,Chenxi;Larsen,KevinP;Hummer,Gerhard;Hanson,PhyllisI;Hurley,JamesH
凋亡相关基因2（Apoptosis linked Gene-2，ALG-2）是一种多功能的细胞内钙离子传感器，是penta-EF手蛋白家族的典型成员。ALG-2在修复质膜和溶酶体膜的损伤中以及在COPII依赖性出芽的atendoplasmicreticulumexitsites（ERES）中起作用。在Ca 2+存在下，ALG-2在膜修复中与ESCRT-1和阿利克斯结合，在ERES处与SEC 31 A结合。ALG-2还通过静电和疏水相互作用的组合在Ca 2+存在下直接结合到酸性膜。通过结合巨大的单层囊泡为基础的实验和分子动力学模拟，我们表明，ALG-2在这些位置的电荷反转突变体破坏膜招聘。ALG-2膜结合突变体响应于Thapsiglavin诱导的Ca 2+释放而减少或消除ERES定位，但在溶酶体Ca 2+释放后仍定位于溶酶体。体外重建表明，ALG-2膜结合缺陷可以通过与ESCRT-I结合来挽救。因此，这些数据揭示了直接Ca 2+依赖性膜结合的性质及其与ALG-2的细胞功能中的Ca 2+依赖性蛋白结合的相互作用。
Apoptosis linked Gene-2 (ALG-2) is a multifunctional intracellular Ca2+sensor and the archetypal member of the penta-EF hand protein family. ALG-2 functions in the repair of damage to both the plasma and lysosome membranes and in COPII-dependent budding atendoplasmicreticulumexitsites (ERES). In the presence of Ca2+, ALG-2 binds to ESCRT-I and ALIX in membrane repair and to SEC31A at ERES. ALG-2 also binds directly to acidic membranes in the presence of Ca2+by a combination of electrostatic and hydrophobic interactions. By combining giant unilamellar vesicle-based experiments and molecular dynamics simulations, we show that charge-reversed mutants of ALG-2 at these locations disrupt membrane recruitment. ALG-2 membrane binding mutants have reduced or abrogated ERES localization in response to Thapsigargin-induced Ca2+release but still localize to lysosomes following lysosomal Ca2+release. In vitro reconstitution shows that the ALG-2 membrane-binding defect can be rescued by binding to ESCRT-I. These data thus reveal the nature of direct Ca2+-dependent membrane binding and its interplay with Ca2+-dependent protein binding in the cellular functions of ALG-2.