Dynamin superfamily molecular motors use guanosine triphosphate (GTP) as a source of energy for membrane-remodeling events. We found that knockdown of nucleoside diphosphate kinases (NDPKs) NM23-H1/H2, which produce GTP through adenosine triphosphate (ATP)-driven conversion of guanosine diphosphate (GDP), inhibited dynamin-mediated endocytosis. NM23-H1/H2 localized at clathrin-coated pits and interacted with the proline-rich domain of dynamin. In vitro, NM23-H1/H2 were recruited to dynamin-induced tubules, stimulated GTP-loading on dynamin, and triggered fission in the presence of ATP and GDP. NM23-H4, a mitochondriaspecific NDPK, colocalized with mitochondrial dynamin-like OPA1 involved in mitochondria * Corresponding author. mathieu.boissan@inserm.fr (M.B.); philippe.chavrier@curie.fr (P.C. Europe PMC Funders Author ManuscriptsEurope PMC Funders Author Manuscripts inner membrane fusion and increased GTP-loading on OPA1. Like OPA1 loss of function, silencing of NM23-H4 but not NM23-H1/H2 resulted in mitochondrial fragmentation, reflecting fusion defects. Thus, NDPKs interact with and provide GTP to dynamins, allowing these motor proteins to work with high thermodynamic efficiency.The 100-kD dynamin guanosine triphosphatase (GTPase) promotes uptake of cell-surface receptors both by clathrin-dependent and -independent pathways (1, 2). Dynamin polymerizes into helix around the neck of endocytic pits and induces guanosine triphosphate (GTP) hydrolysis-driven membrane fission (3-7). Typical of molecular motors, dynamin has a low affinity for GTP and a high basal GTP-hydrolysis rate, which can be further stimulated by dynamin polymerization (8,9). This maximizes chemical energy gain and kinetics of hydrolysis, respectively, which in vivo depend on high concentration ratios of adenosine triphosphate/adenosine diphosphate (ATP/ADP) or GTP/guanosine diphosphate (GDP). The cellular concentrations of GTP and GDP are at least a factor of 10 lower than those of ATP and ADP, and GTP/GDP ratios could thus decrease much more rapidly at elevated workload, both of which make GTP not an ideal substrate for high-turnover, energy-dependent enzymes. Paradoxically, dynamin GTPases are among the most powerful molecular motors described (7).Studies in Drosophila identified a genetic interaction between dynamin and Awd (10-12). Awd belongs to the family of nucleoside diphosphate kinases (NDPKs), which catalyze synthesis of nucleoside triphosphates, including GTP, from corresponding nucleoside diphosphates and ATP (13). The most abundant human NDPKs are the highly related cytosolic proteins NM23-H1 and -H2. NM23-H4, another NDPK-family member, localizes exclusively at the mitochondrial inner membrane (14, 15). Mitochondrial membrane dynamics require dynamin-related GTPases (16). We hypothesized that NDPKs could influence the function of dynamin family members in membrane-remodeling events through spatially controlled GTP production and availability.Knockdown of NM23-H1 and -H2 (fig. S1, A to E) reduced clathrin-dependent endocyt...

Loss of NM23-H1 expression correlates with the degree of metastasis and with unfavorable clinical prognosis in several types of human carcinoma. However, the mechanistic basis for the metastasis suppressor function of NM23-H1 is obscure. We silenced NM23-H1 expression in human hepatoma and colon carcinoma cells and methodologically investigated effects on cell-cell adhesion, migration, invasion, and signaling linked to cancer progression. NM23-H1 silencing disrupted cell-cell adhesion mediated by E-cadherin, resulting in β-catenin nuclear translocation and T-cell factor/lymphoid-enhancing factor-1 transactivation. Further, NM23-H1 silencing promoted cellular scattering, motility, and extracellular matrix invasion by promoting invadopodia formation and upregulating several matrix metalloproteinases (MMP), including membrane type 1 MMP. In contrast, silencing the related NM23-H2 gene was ineffective at promoting invasion. NM23-H1 silencing activated proinvasive signaling pathways involving Rac1, mitogen-activated protein kinases, phosphatidylinositol 3-kinase (PI3K)/Akt, and src kinase. Conversely, NM23-H1 was dispensable for cancer cell proliferation in vitro and liver regeneration in NM23-M1 null mice, instead inducing cellular resistance to chemotherapeutic drugs in vitro. Analysis of NM23-H1 expression in clinical specimens revealed high expression in premalignant lesions (liver cirrhosis and colon adenoma) and the central body of primary liver or colon tumors, but downregulation at the invasive front of tumors. Our findings reveal that NM23-H1 is critical for control of cell-cell adhesion and cell migration at early stages of the invasive program in epithelial cancers, orchestrating a barrier against conversion of in situ carcinoma into invasive malignancy. Cancer Res; 70(19); 7710-22. ©2010 AACR.