RESUMEN
Eukaryotic NMEs/NDP kinases are a family of 10 multifunctional proteins that occur in different cellular compartments and interact with various cellular components (proteins, membranes, and DNA). In contrast to the well-studied Group I NMEs (NME1-4), little is known about the more divergent Group II NMEs (NME5-9). Three recent publications now shed new light on NME6. First, NME6 is a third mitochondrial NME, largely localized in the matrix space, associated with the mitochondrial inner membrane. Second, while its monomeric form is inactive, NME6 gains NDP kinase activity through interaction with mitochondrial RCC1L. This challenges the current notion that mammalian NMEs require the formation of hexamers to become active. The formation of complexes between NME6 and RCC1L, likely heterodimers, seemingly obviates the necessity for hexamer formation, stabilizing a NDP kinase-competent conformation. Third, NME6 is involved in mitochondrial gene maintenance and expression by providing (d)NTPs for replication and transcription (in particular the pyrimidine nucleotides) and by a less characterized mechanism that supports mitoribosome function. This review offers an overview of NME evolution and structure and highlights the new insight into NME6. The new findings position NME6 as the most comprehensively studied protein in NME Group II and may even suggest it as a new paradigm for related family members.
Asunto(s)
Mitocondrias , Humanos , Animales , Mitocondrias/metabolismo , Proteínas Mitocondriales/metabolismo , Proteínas Mitocondriales/genética , Nucleósido Difosfato Quinasas NM23/metabolismo , Nucleósido Difosfato Quinasas NM23/genética , Nucleósido Difosfato Quinasa D/metabolismo , Nucleósido Difosfato Quinasa D/genéticaRESUMEN
Nucleoside diphosphate (NDP) kinases 1 and 2 (NME1/2) are well-characterized enzymes known for their NDP kinase activity. Recently, these enzymes have been shown by independent studies to bind coenzyme A (CoA) or acyl-CoA. These findings suggest a hitherto unknown role for NME1/2 in the regulation of CoA/acyl-CoA-dependent metabolic pathways, in tight correlation with the cellular NTP/NDP ratio. Accordingly, the regulation of NME1/2 functions by CoA/acyl-CoA binding has been described, and additionally, NME1/2 have been shown to control the cellular pathways consuming acetyl-CoA, such as histone acetylation and fatty acid synthesis. NME1/2-controlled histone acetylation in turn mediates an important transcriptional response to metabolic changes, such as those induced following a high-fat diet (HFD). This review discusses the CoA/acyl-CoA-dependent NME1/2 activities and proposes that these enzymes be considered as the first identified carriers of CoA/short-chain acyl-CoAs.
Asunto(s)
Adenosina Trifosfato , Humanos , Animales , Adenosina Trifosfato/metabolismo , Acilcoenzima A/metabolismo , Nucleósido Difosfato Quinasas NM23/metabolismo , Nucleósido Difosfato Quinasas NM23/genética , Nucleósido-Difosfato Quinasa/metabolismo , Nucleósido-Difosfato Quinasa/genética , AcetilaciónRESUMEN
Metastasis is a key determinant in cancer mortality which is often associated with decreased levels of Nm23-H1, a well-established metastasis suppressor. Despite lacking a secretion signal peptide, Nm23-H1 has been reported to be present in the extracellular space and enclosed within extracellular vesicles (EVs). While the presence of Nm23-H1 proteins in EVs released by cancer cells has been observed through proteomics profiling, the role of vesicular Nm23-H1 remains unclear. Here, we investigated the function of vesicular Nm23-H1 using MDA-MB-231 (highly metastatic, low Nm23-H1) and MCF-7 (low/non-metastatic, high Nm23-H1) breast cancer cell models. Our findings confirm that Nm23-H1 is indeed encapsulated within EVs, and its levels can be manipulated through overexpression and knockdown approaches. Functional assays revealed that EVs derived from MDA-MB-231 cells that contained high levels of Nm23-H1 exhibit impaired pro-migratory properties, suggesting that vesicular Nm23-H1 may act as a metastasis suppressor. Furthermore, EVs with increased levels of Nm23-H1 altered the transcript levels of multiple cancer-related genes in recipient cells and stimulated type I interferon signaling through STAT1 phosphorylation. These results suggest the existence of an unconventional signaling pathway mediated by the uptake of EVs enriched with Nm23-H1, which may contribute to the anti-metastatic effect of Nm23-H1 in the tumor microenvironment. Additionally, our study demonstrates that elevated Nm23-H1 levels can impact the abundance of various other proteins encapsulated within breast cancer cell-derived EVs, such as SUSD2 (Sushi Domain Containing 2) which can also modulate metastasis.
Asunto(s)
Neoplasias de la Mama , Movimiento Celular , Vesículas Extracelulares , Nucleósido Difosfato Quinasas NM23 , Humanos , Vesículas Extracelulares/metabolismo , Nucleósido Difosfato Quinasas NM23/metabolismo , Nucleósido Difosfato Quinasas NM23/genética , Neoplasias de la Mama/patología , Neoplasias de la Mama/metabolismo , Femenino , Línea Celular Tumoral , Factor de Transcripción STAT1/metabolismo , Células MCF-7RESUMEN
BACKGROUND: Colorectal cancer (CRC) ranks third in cancer incidence globally and is the second leading cause of cancer-related mortality. The nucleoside diphosphate kinase 1 (NME1) and netrin 1 receptor (DCC) genes have been associated with resistance against tumorigenesis and tumor metastasis. This study investigates the potential association between NME1 (rs34214448 G > T and rs2302254 C > T) and DCC (rs2229080 G > C and rs714 A > G) variants and susceptibility to colorectal cancer development. METHODS: Samples from 232 colorectal cancer patients and 232 healthy blood donors underwent analysis. Variants were identified using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) methodology. Associations were assessed using odds ratios (OR), and the p values were adjusted with Bonferroni test. RESULTS: Individuals carrying the G/T and T/T genotypes for the NME1 rs34214448 variant exhibited a higher susceptibility for develop colorectal cancer (OR = 2.68, 95% CI: 1.76-4.09, P = 0.001 and OR = 2.47, 95% CI: 1.37-4.47, P = 0.001, respectively). These genotypes showed significant associations in patients over 50 years (OR = 2.87, 95% CI: 1.81-4.54, P = 0.001 and OR = 2.99, 95% CI: 1.54-5.79, P = 0.001 respectively) and with early Tumor-Nodule-Metastasis (TNM) stage (P = 0.001), and tumor location in the rectum (P = 0.001). Furthermore, the DCC rs2229080 variant revealed that carriers of the G/C genotype had an increased risk for develop colorectal cancer (OR = 2.00, 95% CI: 1.28-3.11, P = 0.002) and were associated with age over 50 years, sex, and advanced TNM stages (P = 0.001). CONCLUSIONS: These findings suggest that the NME1 rs34214448 and DCC rs2229080 variants play a significant role in colorectal cancer development.
Asunto(s)
Neoplasias Colorrectales , Neoplasias Gástricas , Humanos , Persona de Mediana Edad , Predisposición Genética a la Enfermedad , Polimorfismo de Nucleótido Simple , Genotipo , Neoplasias Gástricas/genética , Neoplasias Colorrectales/epidemiología , Neoplasias Colorrectales/genética , Neoplasias Colorrectales/patología , Estudios de Casos y Controles , Receptor DCC/genética , Nucleósido Difosfato Quinasas NM23/genéticaRESUMEN
Gastric cancer (GC) is one of the most leading cause of malignancies. However, the molecular mechanisms underlying stomach carcinogenesis remain incompletely understood. Dysregulated genetic and epigenetic alternations significantly contribute to GC development. Here, we report that ASH1L and its antisense lncRNA ASH1L-AS1, which are transcribed from the most significant GC-risk signal at 1q22, act as novel oncogenes. The high levels of ASH1L or lncRNA ASH1L-AS1 expression in GC specimens are associated with worse prognosis of patients. In line with this, ASH1L and ASH1L-AS1 are functionally important in promoting GC disease progression. LncRNA ASH1L-AS1 up-regulates ASH1L transcription, increases histone methyltransferase ASH1L expression and elevates genome-wide H3K4me3 modification levels in GC cells. Furthermore, ASH1L-AS1 directly interacts with transcription factor NME1 protein to form the ASH1L-AS1-NME1 ribonucleoprotein, which transcriptionally promotes expression of ASH1L, ASH1L-AS1, KRAS and RAF1, and activates the RAS signaling pathway in GC cells. Taken together, our data demonstrated that the ASH1L-AS1-ASH1L regulatory axis controls histone modification reprogram and activation of the RAS signaling in cancers. Thus, ASH1L-AS1 might be a novel targets of GC therapeutics and diagnosis in the clinic.
Asunto(s)
MicroARNs , ARN Largo no Codificante , Neoplasias Gástricas , Humanos , Neoplasias Gástricas/patología , Línea Celular Tumoral , ARN Largo no Codificante/genética , Transducción de Señal/genética , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Regulación Neoplásica de la Expresión Génica , Proliferación Celular/genética , MicroARNs/genética , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , N-Metiltransferasa de Histona-Lisina/genética , N-Metiltransferasa de Histona-Lisina/metabolismo , Nucleósido Difosfato Quinasas NM23/genéticaRESUMEN
BACKGROUND: NME1 has been exploited as a potential translational target for decades. Substantial efforts have been made to upregulate the expression of NME1 and restore its anti-metastasis function in metastatic cancer. METHODS: Cycloheximide (CHX) chase assay was used to measure the steady-state protein stability of NME1 and HSP90α. The NME1-associating proteins were identified by immunoprecipitation combined with mass spectrometric analysis. Gene knockdown and overexpression were employed to examine the impact of HSP90AA1 on intracellular NME1 degradation. The motility and invasiveness of breast cancer cells were examined in vitro using wound healing and transwell invasion assays. The orthotopic spontaneous metastasis and intra-venous experimental metastasis assays were used to test the formation of metastasis in vivo, respectively. RESULTS: HSP90α interacts with NME1 and increases NME1 lifetime by impeding its ubiquitin-proteasome-mediated degradation. HSP90α overexpression significantly inhibits the metastatic potential of breast cancer cells in vitro and in vivo. A novel cell-permeable peptide, OPT22 successfully mimics the HSP90α function and prolongs the life span of endogenous NME1, resulting in reduced metastasis of breast cancer. CONCLUSION: These results not only reveal a new mechanism of NME1 degradation but also pave the way for the development of new and effective approaches to metastatic cancer therapy.
Asunto(s)
Neoplasias de la Mama , Proteínas de Choque Térmico , Humanos , Femenino , Proteínas de Choque Térmico/metabolismo , Neoplasias de la Mama/patología , Línea Celular Tumoral , Procesamiento Proteico-Postraduccional , Proteínas HSP90 de Choque Térmico/metabolismo , Metástasis de la Neoplasia , Nucleósido Difosfato Quinasas NM23/genéticaRESUMEN
Since the identification of NM23 (now called NME1) as the first metastasis suppressor gene (MSG), a small number of other gene products and non-coding RNAs have been identified that suppress specific parameters of the metastatic cascade, yet which have little or no ability to regulate primary tumor initiation or maintenance. MSG can regulate various pathways or cell biological functions such as those controlling mitogen-activated protein kinase pathway mediators, cell-cell and cell-extracellular matrix protein adhesion, cytoskeletal architecture, G-protein-coupled receptors, apoptosis, and transcriptional complexes. One defining facet of this gene class is that their expression is typically downregulated, not mutated, in metastasis, such that any effective therapeutic intervention would involve their re-expression. This review will address the therapeutic targeting of MSG, once thought to be a daunting task only facilitated by ectopically re-expressing MSG in metastatic cells in vivo. Examples will be cited of attempts to identify actionable oncogenic pathways that might suppress the formation or progression of metastases through the re-expression of specific metastasis suppressors.
Asunto(s)
Genes Supresores de Tumor , Nucleósido Difosfato Quinasas NM23 , Humanos , Nucleósido Difosfato Quinasas NM23/genética , Nucleósido Difosfato Quinasas NM23/metabolismo , Metástasis de la NeoplasiaRESUMEN
Mitochondrial fusion plays an important role in both their structure and function. In this issue, Su et al. (2023. J. Cell Biol.https://doi.org/10.1083/jcb.202301091) report that a nucleoside diphosphate kinase, NME3, facilitates mitochondrial tethering prior to fusion through its direct membrane-binding and hexamerization but not its kinase activity.
Asunto(s)
Difosfatos , Mitocondrias , Nucleósido Difosfato Quinasas NM23 , Mitocondrias/genética , Dinámicas Mitocondriales , Nucleótidos , Fosforilación , Humanos , Nucleósido Difosfato Quinasas NM23/genéticaRESUMEN
Coenzyme A (CoA) is a key cellular metabolite which participates in diverse metabolic pathways, regulation of gene expression and the antioxidant defense mechanism. Human NME1 (hNME1), which is a moonlighting protein, was identified as a major CoA-binding protein. Biochemical studies showed that hNME1 is regulated by CoA through both covalent and non-covalent binding, which leads to a decrease in the hNME1 nucleoside diphosphate kinase (NDPK) activity. In this study, we expanded the knowledge on previous findings by focusing on the non-covalent mode of CoA binding to the hNME1. With X-ray crystallography, we solved the CoA bound structure of hNME1 (hNME1-CoA) and determined the stabilization interactions CoA forms within the nucleotide-binding site of hNME1. A hydrophobic patch stabilizing the CoA adenine ring, while salt bridges and hydrogen bonds stabilizing the phosphate groups of CoA were observed. With molecular dynamics studies, we extended our structural analysis by characterizing the hNME1-CoA structure and elucidating possible orientations of the pantetheine tail, which is absent in the X-ray structure due to its flexibility. Crystallographic studies suggested the involvement of arginine 58 and threonine 94 in mediating specific interactions with CoA. Site-directed mutagenesis and CoA-based affinity purifications showed that arginine 58 mutation to glutamate (R58E) and threonine 94 mutation to aspartate (T94D) prevent hNME1 from binding to CoA. Overall, our results reveal a unique mode by which hNME1 binds CoA, which differs significantly from that of ADP binding: the α- and ß-phosphates of CoA are oriented away from the nucleotide-binding site, while 3'-phosphate faces catalytic histidine 118 (H118). The interactions formed by the CoA adenine ring and phosphate groups contribute to the specific mode of CoA binding to hNME1.
Asunto(s)
Nucleótidos , Treonina , Humanos , Cristalografía por Rayos X , Sitios de Unión , Coenzima A , Arginina , Adenina , Nucleósido Difosfato Quinasas NM23/genéticaRESUMEN
Metastatic progression is regulated by metastasis promoter and suppressor genes. NME1, the prototypic and first described metastasis suppressor gene, encodes a nucleoside diphosphate kinase (NDPK) involved in nucleotide metabolism; two related family members, NME2 and NME4, are also reported as metastasis suppressors. These proteins physically interact with members of the GTPase dynamin family, which have key functions in membrane fission and fusion reactions necessary for endocytosis and mitochondrial dynamics. Evidence supports a model in which NDPKs provide GTP to dynamins to maintain a high local GTP concentration for optimal dynamin function. NME1 and NME2 are cytosolic enzymes that provide GTP to dynamins at the plasma membrane, which drive endocytosis, suggesting that these NMEs are necessary to attenuate signaling by receptors on the cell surface. Disruption of NDPK activity in NME-deficient tumors may thus drive metastasis by prolonging signaling. NME4 is a mitochondrial enzyme that interacts with the dynamin OPA1 at the mitochondria inner membrane to drive inner membrane fusion and maintain a fused mitochondrial network. This function is consistent with the current view that mitochondrial fusion inhibits the metastatic potential of tumor cells whereas mitochondrial fission promotes metastasis progression. The roles of NME family members in dynamin-mediated endocytosis and mitochondrial dynamics and the intimate link between these processes and metastasis provide a new framework to understand the metastasis suppressor functions of NME proteins.
Asunto(s)
Nucleósido Difosfato Quinasas NM23 , Neoplasias , Humanos , Nucleósido Difosfato Quinasas NM23/genética , Nucleósido Difosfato Quinasas NM23/metabolismo , Dinaminas/metabolismo , Neoplasias/patología , Membrana Celular/metabolismo , Guanosina TrifosfatoRESUMEN
BACKGROUND: Colorectal cancer (CRC) has a high mortality rate, and therapeutic approaches to treat these cancers are varied and depend on the metabolic state of the tumour. Profiles of CRC tumours have identified several biomarkers, including microRNAs. microRNA-210 (miR-210) levels are directly correlated with CRC survival. miR-210 expression is higher in metastatic colon cancer cells versus non-metastatic and normal colon epithelium. Therefore, efficient methods to inhibit miR-210 expression in CRC may provide new advances in treatments. METHODS: Expression of miRs was determined in several metastatic and non-metastatic cell lines. miR-210 expression was inhibited using PMIS-miR-210 in transduced cells, which were transplanted into xenograft mice. In separate experiments, CRC tumours were allowed to grow in xenograft mice and treated with therapeutic injections of PMIS-miR-210. Molecular and biochemical experiments identified several new pathways targeted by miR-210 inhibition. RESULTS: miR-210 inhibition can significantly reduce tumour growth of implanted colon cancer cells in xenograft mouse models. The direct administration of PMIS-miR-210 to existing tumours can inhibit tumour growth in both NSG and Foxn1nu/j mouse models and is more efficacious than capecitabine treatments. Tumour cells further transfer the PMIS-miR-210 inhibitor to neighbouring cells by extracellular vesicles to inhibit miR-210 throughout the tumour. miR-210 inhibition activates the cleaved caspase 3 apoptotic pathway to reduce tumour formation. We demonstrate that the long non-coding transcript XIST is regulated by miR-210 correlating with decreased XIST expression in CRC tumours. XIST acts as a competing endogenous RNA for miR-210, which reduces XIST levels and miR-210 inhibition increases XIST transcripts in the nucleus and cytoplasm. The increased expression of NME1 is associated with H3K4me3 and H3K27ac modifications in the NME1 proximal promoter by XIST. CONCLUSION: Direct application of the PMIS-miR-210 inhibitor to growing tumours may be an effective colorectal cancer therapeutic.
Asunto(s)
Neoplasias del Colon , Vesículas Extracelulares , MicroARNs , Nucleósido Difosfato Quinasas NM23 , ARN Largo no Codificante , Animales , Apoptosis/genética , Capecitabina , Caspasa 3 , Neoplasias del Colon/genética , Neoplasias del Colon/metabolismo , Neoplasias del Colon/patología , Neoplasias Colorrectales/genética , Neoplasias Colorrectales/metabolismo , Neoplasias Colorrectales/patología , Vesículas Extracelulares/genética , Vesículas Extracelulares/metabolismo , Vesículas Extracelulares/patología , Humanos , Ratones , MicroARNs/genética , MicroARNs/metabolismo , Nucleósido Difosfato Quinasas NM23/genética , Nucleósido Difosfato Quinasas NM23/metabolismo , ARN Largo no Codificante/genética , ARN Largo no Codificante/metabolismoRESUMEN
Tumor-derived exosomes have documented roles in accelerating the initiation and outgrowth of metastases, as well as in therapy resistance. Little information supports the converse, that exosomes or similar vesicles can suppress metastasis. We investigated the NME1 (Nm23-H1) metastasis suppressor as a candidate for metastasis suppression by extracellular vesicles. Exosomes derived from two cancer cell lines (MDA-MB-231T and MDA-MB-435), when transfected with the NME1 (Nm23-H1) metastasis suppressor, secreted exosomes with NME1 as the predominant constituent. These exosomes entered recipient tumor cells, altered their endocytic patterns in agreement with NME1 function, and suppressed in vitro tumor cell motility and migration compared to exosomes from control transfectants. Proteomic analysis of exosomes revealed multiple differentially expressed proteins that could exert biological functions. Therefore, we also prepared and investigated liposomes, empty or containing partially purified rNME1. rNME1 containing liposomes recapitulated the effects of exosomes from NME1 transfectants in vitro. In an experimental lung metastasis assay the median lung metastases per histologic section was 158 using control liposomes and 15 in the rNME1 liposome group, 90.5% lower than the control liposome group (P = 0.016). The data expand the exosome/liposome field to include metastasis suppressive functions and describe a new translational approach to prevent metastasis.
Asunto(s)
Neoplasias de la Mama , Exosomas , Neoplasias Pulmonares , Nucleósido Difosfato Quinasas NM23 , Línea Celular Tumoral , Femenino , Humanos , Liposomas , Neoplasias Pulmonares/secundario , Nucleósido Difosfato Quinasas NM23/genética , Nucleósido Difosfato Quinasas NM23/metabolismo , Metástasis de la Neoplasia , ProteómicaRESUMEN
BACKGROUND: High expression of NME1 is associated with hepatocellular carcinoma (HCC) progression and poor prognosis. However, there are few reports on the association between NME1 and microRNAs (miRNAs) in HCC progression. OBJECTIVES: To explore miRNAs that regulate NME1 expression in HCC. MATERIAL AND METHODS: Data from the Cancer Genome Atlas (TCGA), Human Protein Atlas (HPA), TargetScan, starBase, and mirDIP were used to analyze the expression pattern of NME1 in HCC tissues, the relationship between NME1 level and the progression of HCC or patient prognosis, miRNAs targeting NME1, and the biological processes that may be regulated by NME1. The regulation of miRNAs to NME1 was assessed using the dual-luciferase reporter assay, quantitative reverse transcription polymerase chain reaction (qRT-PCR) and western blotting. The cell cycle and cell proliferation were detected using propidium iodide (PI) staining and EdU assay, respectively. RESULTS: Highly expressed NME1 in HCC was associated with HCC progression and prognosis. The miR-139-5p and miR-335-5p were weakly expressed in HCC samples and negatively correlated with NME1. The downregulation of miR-139-5p in HCC patients resulted in worse overall survival (OS) and disease-free interval (DFI); however, the level of miR-335-5p was not significantly correlated with OS and DFI in patients with HCC. In vitro experiments verified that the level of miR-139-5p was lower and NME1 expression was higher in HCC cell lines compared to L-02. Moreover, miR-139-5p negatively regulates the expression of NME1 in HCC cell lines. The NME1 may regulate cell cycle, DNA replication, oxidative phosphorylation, and the pentose phosphate pathway. The miR-139-5p inhibited cell proliferation by negatively regulating NME1 expression. CONCLUSIONS: The upregulation of NME1 in HCC indicates a poor prognosis. The NME1 is negatively regulated by miR-139-5p to inhibit cell proliferation.
Asunto(s)
Carcinoma Hepatocelular , Neoplasias Hepáticas , MicroARNs , Nucleósido Difosfato Quinasas NM23 , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/patología , Línea Celular , Línea Celular Tumoral , Movimiento Celular/genética , Proliferación Celular/genética , Regulación Neoplásica de la Expresión Génica , Humanos , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patología , MicroARNs/genética , MicroARNs/metabolismo , Nucleósido Difosfato Quinasas NM23/genética , Nucleósido Difosfato Quinasas NM23/metabolismoRESUMEN
Cancer stem cells (CSCs), which can self-renew and produce heterogeneous cancer cells, are the key factors during tumorigenesis. Transcription factors take essential effects on CSCs. However, the role of transcription factors in regulating the stemness of gastric cancer stem-like cells has not been well explored. In this investigation, it was found that transcription factor NME2 (NME/NM23 nucleoside diphosphate kinase 2) was upregulated in gastric cancer stem-like cells that sorted from the solid tumors of patients with gastric cancer and gastric cancer cell lines. NME2 could preserve the stemness of gastric cancer stem-like cells via suppressing their apoptosis. In vitro and in vivo data revealed that NME2 was crucial for maintaining the stemness of gastric cancer stem cells by enhancing the expression of anti-apoptosis genes. Consequently, our data contributed a new perspective to the relationship between transcription factor and the stemness maintenance of gastric cancer stem cells.
Asunto(s)
Nucleósido Difosfato Quinasas NM23/metabolismo , Células Madre Neoplásicas/metabolismo , Neoplasias Gástricas/metabolismo , Neoplasias Gástricas/patología , Factores de Transcripción/metabolismo , Animales , Apoptosis/genética , Secuencia de Bases , Carcinogénesis/genética , Carcinogénesis/patología , Línea Celular Tumoral , Regulación Neoplásica de la Expresión Génica , Silenciador del Gen , Humanos , Ratones Endogámicos NOD , Ratones SCID , Modelos Biológicos , Nucleósido Difosfato Quinasas NM23/genética , Células Madre Neoplásicas/patología , Regulación hacia Arriba/genéticaRESUMEN
BACKGROUND: Mitochondrial nucleoside diphosphate kinase (NDPK-D, NME4, NM23-H4) is a multifunctional enzyme mainly localized in the intermembrane space, bound to the inner membrane. RESULTS: We constructed loss-of-function mutants of NDPK-D, lacking either NDP kinase activity or membrane interaction and expressed mutants or wild-type protein in cancer cells. In a complementary approach, we performed depletion of NDPK-D by RNA interference. Both loss-of-function mutations and NDPK-D depletion promoted epithelial-mesenchymal transition and increased migratory and invasive potential. Immunocompromised mice developed more metastases when injected with cells expressing mutant NDPK-D as compared to wild-type. This metastatic reprogramming is a consequence of mitochondrial alterations, including fragmentation and loss of mitochondria, a metabolic switch from respiration to glycolysis, increased ROS generation, and further metabolic changes in mitochondria, all of which can trigger pro-metastatic protein expression and signaling cascades. In human cancer, NME4 expression is negatively associated with markers of epithelial-mesenchymal transition and tumor aggressiveness and a good prognosis factor for beneficial clinical outcome. CONCLUSIONS: These data demonstrate NME4 as a novel metastasis suppressor gene, the first localizing to mitochondria, pointing to a role of mitochondria in metastatic dissemination.
Asunto(s)
Neoplasias , Nucleósido-Difosfato Quinasa , Animales , Membranas Intracelulares , Ratones , Mitocondrias , Nucleósido Difosfato Quinasas NM23/genética , Nucleósido Difosfato Quinasas NM23/metabolismo , Neoplasias/genética , Neoplasias/metabolismo , Nucleósido Difosfato Quinasa D/metabolismo , Nucleósido-Difosfato Quinasa/genética , Nucleósido-Difosfato Quinasa/metabolismoRESUMEN
Hepatocyte growth factor-overexpressing mice that harbor a deletion of the Ink4a/p16 locus (HP mice) form melanomas with low metastatic potential in response to UV irradiation. Here we report that these tumors become highly metastatic following hemizygous deletion of the Nme1 and Nme2 metastasis suppressor genes (HPN mice). Whole-genome sequencing of melanomas from HPN mice revealed a striking increase in lung metastatic activity that is associated with missense mutations in eight signature genes (Arhgap35, Atp8b4, Brca1, Ift172, Kif21b, Nckap5, Pcdha2, and Zfp869). RNA-seq analysis of transcriptomes from HP and HPN primary melanomas identified a 32-gene signature (HPN lung metastasis signature) for which decreased expression is strongly associated with lung metastatic potential. Analysis of transcriptome data from The Cancer Genome Atlas revealed expression profiles of these genes that predict improved survival of patients with cutaneous or uveal melanoma. Silencing of three representative HPN lung metastasis signature genes (ARRDC3, NYNRIN, RND3) in human melanoma cells resulted in increased invasive activity, consistent with roles for these genes as mediators of the metastasis suppressor function of NME1 and NME2. In conclusion, our studies have identified a family of genes that mediate suppression of melanoma lung metastasis, and which may serve as prognostic markers and/or therapeutic targets for clinical management of metastatic melanoma.
Asunto(s)
Biomarcadores de Tumor/genética , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/secundario , Melanoma/genética , Nucleósido Difosfato Quinasas NM23/genética , Rayos Ultravioleta/efectos adversos , Animales , Perfilación de la Expresión Génica , Regulación Neoplásica de la Expresión Génica , Factor de Crecimiento de Hepatocito/genética , Humanos , Melanoma/etiología , Ratones , Mutación Missense , Análisis de Secuencia de ARN , Análisis de Supervivencia , Secuenciación Completa del GenomaRESUMEN
MicroRNAs (miRNAs) are small noncoding RNAs that function as critical posttranscriptional regulators in various biological processes. While most miRNAs are generated from processing of long primary transcripts via sequential Drosha and Dicer cleavage, some miRNAs that bypass Drosha cleavage can be transcribed as part of another small noncoding RNA. Here, we develop the target-oriented miRNA discovery (TOMiD) bioinformatic analysis method to identify Drosha-independent miRNAs from Argonaute crosslinking and sequencing of hybrids (Ago-CLASH) data sets. Using this technique, we discovered a novel miRNA derived from a primate specific noncoding RNA, the small NF90 associated RNA A (snaR-A). The miRNA derived from snaR-A (miR-snaR) arises independently of Drosha processing but requires Exportin-5 and Dicer for biogenesis. We identify that miR-snaR is concurrently up-regulated with the full snaR-A transcript in cancer cells. Functionally, miR-snaR associates with Ago proteins and targets NME1, a key metastasis inhibitor, contributing to snaR-A's role in promoting cancer cell migration. Our findings suggest a functional link between a novel miRNA and its precursor noncoding RNA.
Asunto(s)
Biología Computacional/métodos , MicroARNs/metabolismo , ARN Largo no Codificante/genética , Regiones no Traducidas 3' , Movimiento Celular , ARN Helicasas DEAD-box/metabolismo , Células HCT116 , Células HEK293 , Humanos , Carioferinas/metabolismo , Células MCF-7 , Nucleósido Difosfato Quinasas NM23/antagonistas & inhibidores , Nucleósido Difosfato Quinasas NM23/genética , Neoplasias/patología , ARN Largo no Codificante/metabolismo , Ribonucleasa III/metabolismoRESUMEN
PURPOSE: To explore the influence of nm23-H1 on the proliferation and apoptosis of glioma cells and the mechanism of action. METHODS: The changes in the messenger RNA (mRNA) expression of nm23-H1 were detected via quantitative real-time-polymerase chain reaction (qRT-PCR), and the relative protein expression level of nm23-H1 was determined using immunohistochemistry. The glioma H4 cells were transfected exogenously with nm23-H1 gene (nm23-H1 group) or empty vector (Vector group), and the biological influence of the expression level of nm23-H1 on H4 cells was then assessed through in vitro functional experiments. Besides, the cells transfected with nm23-H1 were incubated with the protein kinase C (PKC) pathway inhibitor Calphostin C, and functional experiments were performed to observe the changes in the proliferation and apoptosis of cells after incubation. RESULTS: According to the immunohistochemistry and qRT-PCR results, the protein and mRNA expression levels of nm23-H1 declined notably in glioma tissues (p<0.01). The cells with up-regulated nm23-H1 expression had substantially weakened proliferation and migration abilities, but exhibited dramatically enhanced apoptosis (p<0.01). The PKC pathway inhibitor considerably potentiated the effects of nm23-H1 protein on the proliferation and apoptosis of H4 cells (p<0.05), and the protein expression level of nm23-H1 rose in the cells treated with the PKC inhibitor (p<0.01). CONCLUSIONS: Compared with normal brain tissues, nm23-H1 is lowly expressed in glioma tissues and affects the expression of PKC to influence the biological behaviors of H4 cells.
Asunto(s)
Neoplasias Encefálicas/metabolismo , Glioma/metabolismo , Nucleósido Difosfato Quinasas NM23/metabolismo , Proteína Quinasa C/metabolismo , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/patología , Proliferación Celular/fisiología , Glioma/genética , Glioma/patología , Humanos , Nucleósido Difosfato Quinasas NM23/genética , Transducción de SeñalRESUMEN
Tumor metastasis remains an obstacle in cancer treatment and is responsible for most cancer-related deaths. Nm23-H1 is one of the first metastasis suppressor proteins discovered with the ability to inhibit metastasis of many cancers including breast, colon, and liver cancer. Although loss of Nm23-H1 is observed in aggressive cancers and correlated with metastatic potential, little is known regarding the mechanisms that regulate its cellular level. Here, we examined the mechanisms that control Nm23-H1 expression in breast cancer cells. Initial studies in aggressive MDA-MB-231 cells (expressing low Nm23-H1) and less invasive MCF-7 cells (expressing high Nm23-H1) revealed that mRNA levels correlated with protein expression, suggesting that transcriptional mechanisms may control Nm23-H1 expression. Truncational analysis of the Nm23-H1 promoter revealed a proximal and minimal promoter that harbor putative binding sites for transcription factors including CTCF and EGR1. CTCF and EGR1 induced Nm23-H1 expression and reduced cell migration of MDA-MB-231 cells. Moreover, CTCF and EGR1 were recruited to the Nm23-H1 promoter in MCF-7 cells and their expression correlated with Nm23-H1 levels. This study indicates that loss of Nm23-H1 in aggressive breast cancer is apparently caused by downregulation of CTCF and EGR1, which potentially drive Nm23-H1 expression to promote a less invasive phenotype.
Asunto(s)
Biomarcadores de Tumor/metabolismo , Neoplasias de la Mama/patología , Factor de Unión a CCCTC/metabolismo , Movimiento Celular , Proteína 1 de la Respuesta de Crecimiento Precoz/metabolismo , Regulación Neoplásica de la Expresión Génica , Nucleósido Difosfato Quinasas NM23/genética , Apoptosis , Biomarcadores de Tumor/genética , Neoplasias de la Mama/genética , Neoplasias de la Mama/metabolismo , Factor de Unión a CCCTC/genética , Proliferación Celular , Proteína 1 de la Respuesta de Crecimiento Precoz/genética , Femenino , Humanos , Nucleósido Difosfato Quinasas NM23/metabolismo , Células Tumorales CultivadasRESUMEN
BACKGROUND: The aim of the present work was to evaluate the prognostic significance in patients with T1 breast cancer of tissue expression of the two oncosuppressors phosphatase and tensin homolog (PTEN) and non-metastatic clone 23 (NM23) as detected by immunohistochemistry. MATERIALS AND METHODS: We prospectively analyzed 62 patients who underwent surgery for a T1 stage breast cancer. Expression of PTEN and NM23 was tested for correlation with clinical characteristics and clinical outcome. RESULTS: Of the 62 patients considered for our study, 16 underwent mastectomy and 46 underwent conservative surgical treatment. The surgery was considered radical (R0) in all cases described. PTEN and NM23 expression was higher in patients with no lymph node metastases and no recurrent cancer at a mean follow-up of 36 months (range=6-48 months). This correlation was more evident when both PTNE and NM23 expression were highly expressed (p<0.0001). CONCLUSION: Low or lack of PTEN and NM23 immunohistochemical expression in cancer tissue is a risk factor for lymph node involvement and recurrent disease. It may represent a valid prognostic factor in planning therapy in patients who had surgery for T1 breast cancer.