The Nucleoside Diphosphate Kinase D (NM23-H4) Binds the Inner Mitochondrial Membrane with High Affinity to Cardiolipin and Couples Nucleotide Transfer with Respiration

Abstract

Nucleoside diphosphate kinases (NDPK), 3 encoded by NME genes (also called NM23), catalyze the exchange of ␥-phosphate between di-and triphosphonucleosides and participate in the regulation of intracellular nucleotide homeostasis. They mainly utilize ATP formed by oxidative phosphorylation to synthesize the other triphosphonucleosides, in particular GTP (1). Given the poor substrate selectivity of NDPKs, it is assumed that specificity could arise from the presence of different isoforms at different subcellular localizations. Associated in networks with other nucleotide-metabolizing enzymes such as adenylate kinases, creatine kinases, and glycolytic enzymes, NDPKs participate in high energy phosphoryl transfer and signal communication in the cell (2). Up to now nine genes encoding NDPK or NDPK-like proteins have been identified (3, 4), but little is known about their respective role within the cell. The most studied, NDPK-A and -B, encoded by NME1 and NME2 genes, respectively, play a key role in tumor progression and metastasis dissemination (5, 6).NDPK activity has been found associated with different cellular compartments, such as cytosol, nucleus, plasma membrane, and mitochondria. Precise localization in the latter organelles has been a matter of debate. Depending on species and tissue examined, NDPK activity was reported in both the matrix and the intermembrane/cristae space (7), including the so-called contact sites between inner and outer membrane (8 -10). In mammalian liver (rat and rabbit), the NDPK activity was mainly associated with an extra-matrix compartment, probably the intermembrane/cristae space, whereas in heart activity was more abundant in the matrix (11). For mitochondrial NDPK in matrix, many functions have been proposed ranging from nucleotide supply for mitochondrial nucleic acid and protein synthesis to functional interaction with the Krebs * This work was supported by the Germaine de Stael Program for FrancoSwiss collaboration (to U. S. and M.-L. L.), the Agence Nationale de la Recherche (Chaire d'Excellence (to U. S.)), the Marie Curie Intraeuropean Fellowship of the European Community (to M. T.-S.), INSERM, and grants from the Groupement des Entreprises Françaises contre le Cancer and from the Association pour la Recherche contre le Cancer (to M.-L. L.). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

NDPK-D (NM23-H4)-mediated externalization of cardiolipin enables elimination of depolarized mitochondria by mitophagy

“…4,13 HeLa cells constitutively express negligible levels of NDPK-D, 14 which allowed us to introduce either w/t or mutant protein.…”

“…In vitro, the kinase complex shows intermembrane lipid transfer activity as soon as it can simultaneously bind to two membranes containing some CL. 15 However, NDPK-D interacts not only with CL, 14 but also with OPA1, 7 a dynamin-like GTPase. OPA1, beyond its classical role in inner membrane fusion, 41,42 is also directly implicated in mitochondrial quality control via its stressregulated proteolytic cleavage via OMA1.…”

“…The pcDNA4/TO vectors expressing w/t NDPK-D and R90D mutant were prepared as previously described. 14 For determination of mitochondrial volume/fragmentation changes, cells were first infected with adenoviral vector expressing mitochondria directed mCerulean/mFAP prior to treatment. Cells were then incubated with 5 nM malachite green to reveal the transgene prior to imaging (thereby defining the mitochondria 58 ) and mounted in a Tokai Hit environmental stage (Tokyo, Japan) in a Nikon TI inverted microscope equipped with a sweptfield confocal head.…”

NME4/nucleoside diphosphate kinase D in cardiolipin signaling and mitophagy

“…Besides its NDP kinase activity, it binds to both mitochondrial membranes via CL and other anionic phospholipids to induce contact sites. We have shown first for reconstituted in vitro systems, 43 and later for mitochondria and living cells 44,45 that NME4/NDPK-D can also support intermembrane phospholipid transfer. We found in different cellular model systems that a collapse of ΔΨ (eg, using the protonophoric uncoupler CCCP) is triggering CL externalization and induction of mitophagy, and that this requires the presence of CL-binding competent NME4/NDPK-D that exposes triads of basic amino acids.…”

Nucleoside diphosphate kinases (NDPKs) in animal development

“…These multiple functions assigned to the NM23 family of proteins have thus far been difficult to reconcile because the exact cellular function of NM23 has remained unclear. However, beyond nucleoside diphosphate kinase activity several other molecular activities have been linked to NDPKs, such as histidine-dependent protein kinase (histidine phosphotransferase) [18,19] and nuclease activity [20,21], as well as lipid bilayer binding [22][23][24][25][26].…”