A molecular triage process mediated by RING finger protein 126 and BCL2-associated athanogene 6 regulates degradation of G0/G1 switch gene 2
A molecular triage process mediated by RING finger protein 126 and BCL2-associated athanogene 6 regulates degradation of G0/G1 switch gene 2
复制标题
DOI：
10.1074/jbc.ra119.008544
复制
发表时间：
2019-10-04
影响因子：
4.8
通讯作者：
Takashima, Seiji
中科院分区：
文献类型：
--
作者：
Kamikubo, Kenta;Kato, Hisakazu;Takashima, Seiji
在呼吸细胞中，氧化磷酸化产生了大部分ATP。ATP是一种必需的能量来源，特别是在心肌细胞中，因为它们持续收缩和舒张。此前，我们报道了G(0)/G(1)开关基因2 （G0S2）通过与FOF1-ATP合成酶相互作用，正调控线粒体ATP的产生。G0S2过表达可减轻心肌细胞缺血时ATP的下降，并显著增加心肌细胞的缺氧耐受性。在这里，我们发现G0S2蛋白通过细胞质分子分类系统进行蛋白酶体降解，抑制这一过程会增加缺氧时线粒体ATP的产生。首先，我们对针对泛素相关基因的sirna文库进行了筛选，并鉴定出RING finger protein 126 （RNF126）是参与G0S2降解的E3连接酶。rnf126缺陷细胞表现出G0S2蛋白周转延长和G0S2泛素化降低。参与新生膜蛋白分子分选的bcl2相关无thanogene 6 （BAG6）在体外和体内均增强了rnf126介导的G0S2泛素化。接下来，我们发现G0S2疏水区的Glu-44是G0S2多泛素化和蛋白酶体降解所必需的降解物。由于这种降解是G0S2与BAG6相互作用所必需的，因此替代丙氨酸的G0S2突变体（E44A）逃脱了降解。在原代培养的心肌细胞中，G0S2 E44A突变体的过表达和RNF126的敲低都有效地减弱了缺氧条件下ATP的下降。我们得出结论，RNF126/BAG6复合物有助于G0S2的降解，防止G0S2降解的干预措施可能为管理缺血性疾病提供治疗策略。
Oxidative phosphorylation generates most of the ATP in respiring cells. ATP is an essential energy source, especially in cardiomyocytes because of their continuous contraction and relaxation. Previously, we reported that G(0)/G(1) switch gene 2 (G0S2) positively regulates mitochondrial ATP production by interacting with FOF1-ATP synthase. G0S2 overexpression mitigates ATP decline in cardiomyocytes and strongly increases their hypoxic tolerance during ischemia. Here, we show that G0S2 protein undergoes proteasomal degradation via a cytosolic molecular triage system and that inhibiting this process increases mitochondrial ATP production in hypoxia. First, we performed screening with a library of siRNAs targeting ubiquitin-related genes and identified RING finger protein 126 (RNF126) as an E3 ligase involved in G0S2 degradation. RNF126-deficient cells exhibited prolonged G0S2 protein turnover and reduced G0S2 ubiquitination. BCL2-associated athanogene 6 (BAG6), involved in the molecular triage of nascent membrane proteins, enhanced RNF126-mediated G0S2 ubiquitination both in vitro and in vivo. Next, we found that Glu-44 in the hydrophobic region of G0S2 acts as a degron necessary for G0S2 polyubiquitination and proteasomal degradation. Because this degron was required for an interaction of G0S2 with BAG6, an alanine-replaced G0S2 mutant (E44A) escaped degradation. In primary cultured cardiomyocytes, both overexpression of the G0S2 E44A mutant and RNF126 knockdown effectively attenuated ATP decline under hypoxic conditions. We conclude that the RNF126/BAG6 complex contributes to G0S2 degradation and that interventions to prevent G0S2 degradation may offer a therapeutic strategy for managing ischemic diseases.