NME3 is a gatekeeper for DRP1-dependent mitophagy in hypoxia

NME3 is a member of the nucleoside diphosphate kinase (NDPK) family localized on the mitochondrial outer membrane (MOM). Here, we report a role of NME3 in hypoxia-induced mitophagy dependent on its active site phosphohistidine but not the NDPK function. Mice carrying a knock-in mutation in the Nme3 gene disrupting NME3 active site histidine phosphorylation are vulnerable to ischemia/reperfusion-induced infarction and develop abnormalities in cerebellar function. Our mechanistic analysis reveals that hypoxia-induced phosphatidic acid (PA) on mitochondria is essential for mitophagy and the interaction of DRP1 with NME3. The PA binding function of MOM-localized NME3 is required for hypoxia-induced mitophagy. Further investigation demonstrates that the interaction with active NME3 prevents DRP1 susceptibility to MUL1-mediated ubiquitination, thereby allowing a sufficient amount of active DRP1 to mediate mitophagy. Furthermore, MUL1 overexpression suppresses hypoxia-induced mitophagy, which is reversed by co-expression of ubiquitin-resistant DRP1 mutant or histidine phosphorylatable NME3. Thus, the site-specific interaction with active NME3 provides DRP1 a microenvironment for stabilization to proceed the segregation process in mitophagy.

NME3属于核苷二磷酸激酶（NDPK）家族，定位于线粒体外膜（MOM）。本研究报道了NME3在缺氧诱导的线粒体自噬中的作用，该作用依赖于其活性位点的磷酸化组氨酸，而非NDPK的催化功能。在Nme3基因中引入敲入突变以破坏NME3活性位点组氨酸磷酸化的小鼠，更易发生缺血/再灌注诱导的脑梗死，并出现小脑功能异常。本研究的机制分析显示，缺氧诱导的线粒体内磷脂酸（PA）生成对于线粒体自噬以及DRP1与NME3的相互作用至关重要。定位于线粒体外膜的NME3所具备的PA结合功能，是缺氧诱导线粒体自噬所必需的。进一步研究表明，与活性形式NME3的相互作用可避免DRP1被MUL1介导的泛素化修饰，从而使足量的活性DRP1能够介导线粒体自噬过程。此外，MUL1过表达会抑制缺氧诱导的线粒体自噬，而共表达泛素抗性DRP1突变体或可发生组氨酸磷酸化的NME3，可逆转这一抑制效应。综上，与活性形式NME3的位点特异性相互作用，为DRP1提供了稳定化的微环境，以推进线粒体自噬中的分离过程。