| 2000 |
NME4 (Nm23-H4) encodes a mitochondrial nucleoside diphosphate kinase; the full-length protein is inactive due to its N-terminal mitochondrial targeting extension, while the truncated form lacking the extension possesses NDP kinase activity. Import into mitochondria is accompanied by cleavage of the N-terminal extension, restoring activity. X-ray crystallography confirmed the protein forms a hexamer, and submito-chondrial fractionation showed it is associated with mitochondrial membranes, possibly at contact sites between outer and inner membranes. |
Recombinant protein expression in E. coli, NDP kinase activity assay, X-ray crystallography, site-directed mutagenesis (S129P), GFP-fusion confocal microscopy, Western blot subcellular fractionation in HEK293 cells |
The Journal of biological chemistry |
High |
10799505
|
| 2008 |
NME4/NDPK-D binds the inner mitochondrial membrane primarily through electrostatic interaction with cardiolipin (the anionic phospholipid most enriched in the inner membrane), mediated by a surface-exposed basic RRK motif (Arg-90). Mutation R90D strongly reduces phospholipid binding in vitro and in vivo. The membrane-bound state of NME4 is required for functional coupling with oxidative phosphorylation (respiration stimulated by TDP only in mitochondria expressing wild-type, not R90D, NME4). NME4's symmetrical hexameric structure allows it to cross-link anionic phospholipid-containing liposomes, suggesting a role in promoting intermembrane contacts. |
Surface plasmon resonance with recombinant protein and model liposomes, site-directed mutagenesis (R90D), stable expression in HeLa cells, respiration assays, latency assays with isolated mitochondria, antibody binding to mitoplasts |
The Journal of biological chemistry |
High |
18635542
|
| 2012 |
NME4 has a dual function acting as a mitochondrial switch: (1) phosphotransfer/NDP kinase activity supplying GTP locally, and (2) selective intermembrane cardiolipin transfer from inner to outer mitochondrial membrane. Cardiolipin binding inhibits NDP kinase activity but is required for lipid transfer. Wild-type NME4 (but not a membrane-binding-deficient mutant) selectively increased cardiolipin content in the outer mitochondrial membrane. NME4 forms a complex with the mitochondrial GTPase OPA1 in rat liver, suggesting direct local GTP delivery. Wild-type NME4-expressing HeLa cells showed increased Bax accumulation in mitochondria and were sensitized to rotenone-induced apoptosis (cytochrome c release, caspase 3/7 activation, annexin V binding). |
Co-immunoprecipitation (NME4-OPA1 complex), LC-MS lipid analysis of HeLa cells expressing wild-type vs. membrane-binding-deficient mutant NME4, apoptosis assays (cytochrome c release, caspase 3/7, annexin V), molecular modeling |
The Journal of biological chemistry |
High |
23150663
|
| 2016 |
NME4/NDPK-D facilitates translocation of cardiolipin from the inner mitochondrial membrane to the outer mitochondrial membrane surface upon mitophagy induction (CCCP treatment), enabling cardiolipin to serve as an 'eat-me' signal recognized by LC3. RNAi knockdown of NME4 decreased CCCP-induced CL externalization and mitochondrial degradation. The CL-binding deficient mutant R90D was inactive in promoting mitophagy. Proximity ligation assay showed NME4's CL-transfer activity is closely associated with the dynamin-like GTPase OPA1, implicating fission-fusion dynamics. NME4 knockdown also suppressed rotenone- and 6-hydroxydopamine-triggered mitophagy in SH-SY5Y cells. |
RNAi knockdown, CCCP/rotenone/6-OHDA-induced mitophagy, CL externalization assay, R90D mutant functional analysis, in situ proximity ligation assay (PLA) for NME4-OPA1 association, mitochondrial degradation assays in MLE-12, HeLa, and SH-SY5Y cells |
Cell death and differentiation |
High |
26742431
|
| 2015 |
NME4/NDPK-D is acetylated, and its acetylation state regulates its subcellular localization between nucleus and cytoplasm, as well as cell survival. SIRT1 was identified as a binding partner of NME4 by yeast two-hybrid screening, confirmed by co-immunoprecipitation. SIRT1 inhibition increases NME4 acetylation. Overexpression of NME4 with SIRT1, or mutation of acetylated lysine residues in NME4, increases nuclear accumulation. Acetylation-mimic mutant NME4 increased apoptosis in N1E-115 cells. NME4 knockdown induced apoptosis in neuroblastoma cells and mouse cortex. |
Yeast two-hybrid screening, co-immunoprecipitation, site-directed mutagenesis (acetylation-mimic), SIRT1 inhibitor treatment, confocal microscopy, apoptosis assays, in vivo knockdown in mouse cortex |
PloS one |
Medium |
26426123
|
| 2014 |
NME4 suppresses cell migration and invasion in oral cancer through the NME4-JNK-TIMP1-MMP signaling pathway; miR-196 inhibits NME4 expression, thereby activating p-JNK, suppressing TIMP1, and augmenting MMP1/9, promoting invasive phenotype. |
miR-196 overexpression/inhibition, RT-qPCR, Western blot, luciferase reporter assay for miR-196 targeting of NME4 3'UTR, cell migration and invasion assays, confocal microscopy |
Molecular cancer |
Medium |
25233933
|
| 2021 |
NME4 acts as a metastasis suppressor in cancer cells. Loss-of-function mutations (lacking either NDP kinase activity or membrane interaction) or RNAi depletion of NME4 promoted epithelial-mesenchymal transition, increased migratory and invasive potential, and increased metastasis formation in immunocompromised mice. Mechanistically, NME4 loss caused mitochondrial fragmentation and loss, metabolic switch from respiration to glycolysis, and increased ROS generation, triggering pro-metastatic signaling cascades. |
Loss-of-function mutants (kinase-dead and membrane interaction-deficient), RNAi knockdown, in vitro migration/invasion assays, EMT marker analysis, in vivo xenograft metastasis model in immunocompromised mice, metabolic profiling |
BMC biology |
High |
34674701
|
| 2023 |
NME4 interacts with key enzymes in coenzyme A (CoA) metabolism and increases the levels of acetyl-CoA and malonyl-CoA in the liver, promoting triglyceride accumulation and NAFLD progression. Hepatic deletion of Nme4 in mice suppressed hepatic steatosis progression. |
Hepatic-specific Nme4 knockout in mice (high-fat diet model), proteomics, metabolomics, CoA metabolite measurements |
EMBO reports |
Medium |
38177901
|
| 2024 |
NME4 negatively regulates the NFκB2-CCL5 signaling axis in esophageal squamous cell carcinoma, preventing CD8+ T cell infiltration into the tumor microenvironment. Mechanistically, NME4 suppresses NFκB2 activity, which controls CCL5 chemokine expression. |
Syngeneic tumor model in C57BL/6 mice, single-cell RNA sequencing, quantitative proteomics, protein microarray screening, NME4 modulation in murine ESCC cell line AKR |
Immunology |
Medium |
39016535
|
| 2021 |
Let-7f-5p miRNA directly targets the 3' UTR of Nme4 mRNA and negatively regulates Nme4 expression in mouse bone marrow-derived mesenchymal stem cells (BM-MSCs). TNF-α upregulates let-7f-5p (via NF-κB), reducing Nme4 and impairing osteogenic differentiation. Ectopic Nme4 expression reversed the inhibitory effects of let-7f-5p on osteogenesis in vitro and restored bone formation in ovariectomized mice in vivo. |
miRNA mimic/inhibitor transfection, luciferase reporter assay (let-7f-5p targeting Nme4 3'UTR), NME4 overexpression rescue, in vivo ovariectomized mouse model, osteogenic differentiation assays (ALP, Alizarin Red staining) |
Biochemistry and cell biology |
Medium |
34297624
|
| 2026 |
E3 ubiquitin ligase RNF6 directly binds NME4 and promotes its K48-linked polyubiquitination, leading to proteasomal degradation of NME4. NME4 degradation by RNF6 activates the JNK/c-JUN signaling pathway, promoting ovarian cancer malignancy. |
Co-immunoprecipitation (RNF6-NME4 interaction), cycloheximide chase assay, ubiquitination assay (K48-linked), RNF6/NME4 co-modulation rescue experiments, in vivo nude mouse xenograft model |
Pathology, research and practice |
Medium |
41616518
|
| 2024 |
Conserved Arg27 across group I NDPKs (NME1-4) is a key residue for hexamer assembly; Arg27 mutation leads to decreased binding affinity, altered dynamics, and complex destabilization. For NME4 specifically, double and triple Arg mutations destabilize the hexamer into a dimer, partly due to its shorter C-terminal region. |
Molecular dynamics simulations, structural modeling, binding affinity calculations with mutant NME4 constructs |
bioRxivpreprint |
Low |
bio_10.1101_2024.09.19.613900
|