RESUMEN
Rationale: Fibrosis is a pathologic condition of abnormal accumulation of collagen fibrils. Collagen is a major extracellular matrix (ECM) protein synthesized and secreted by myofibroblasts, composing mainly (Gly-X-Y)n triplet repeats with >30% Gly residue. During fibrosis progression, myofibroblasts must upregulate glycine metabolism to meet the high demands of amino acids for collagen synthesis. Method: Expression of PKM2 in myofibroblasts was analyzed in cultured fibroblasts and fibrosis disease tissues. Functional roles of PKM2 and PKM2 activator in biosynthesis of serine â glycine and production of collagen from glycolysis intermediates were assayed in cultured activated LX-2 and human primary lung fibroblast cells. Mouse models of Liver, lung, and pancreas fibrosis were employed to analyze treatment effects of PKM2 activator in organ tissue fibrosis. Results: We report here that myofibroblast differentiation upregulates pyruvate kinase M2 (PKM2) and promotes dimerization of PKM2. Dimer PKM2 slows the flow rate of glycolysis and channels glycolytic intermediates to de novo glycine synthesis, which facilitates collagen synthesis and secretion in myofibroblasts. Our results show that PKM2 activator that converts PKM2 dimer to tetramer, inhibits fibrosis progression in mouse models of liver, lung, and pancreatic fibrosis. Furthermore, metabolism alteration by dimer PKM2 increases NADPH production, which consequently protects myofibroblasts from apoptosis. Conclusion: Our study uncovers a novel role of PKM2 in tissue/organ fibrosis, suggesting a possible strategy for treatment of fibrotic diseases using PKM2 activator.
Asunto(s)
Fibrosis/metabolismo , Glicina/metabolismo , Piruvato Quinasa/metabolismo , Animales , Apoptosis , Diferenciación Celular , Colágeno/metabolismo , Matriz Extracelular/metabolismo , Femenino , Fibroblastos/metabolismo , Fibrosis/fisiopatología , Glicina/fisiología , Glucólisis/efectos de los fármacos , Humanos , Hígado/patología , Pulmón/patología , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Miofibroblastos/metabolismo , Miofibroblastos/fisiología , Páncreas/patología , Piruvato Quinasa/fisiología , Transducción de SeñalRESUMEN
The Pyruvate kinase isozymes M2 (PKM2) protein is a metabolic enzyme that regulates the final step of glycolysis. This enzyme is present in highly proliferating cells and is expressed in the placenta. We recently demonstrated upregulated placental PKM2 during human intrauterine growth restriction (IUGR). Our current objective was to determine PKM2 regulation of trophoblast invasion, trophoblast PKM2 localization as well as mTOR protein expression, and to determine effects of activation of PKM2 during IUGR. Human placental tissues were obtained and analyzed by immunohistochemistry and western blot. Trophoblast cells were cultured in normoxic and hypoxic conditions and real time cell invasion and PKM2 protein were determined during activation (Fructose-6-bisphosphate; FBP6) or inhibition (Shikonin) of PKM2. In vivo studies determined the effects of PKM2 activation on placental and fetal weights. IUGR samples had elevated levels of p-PKM2. Different trophoblast PKM2 localization and expression was observed during normoxia and hypoxia. Decreased trophoblast invasion and PKM2 expression was observed during mTOR inhibition. Protection from decreased placental and fetal weights was observed by PKM2 activation. We conclude that PKM2 regulates trophoblast cell invasion depending on its subcellular location. Our results suggest that PKM2 regulation in trophoblast cells is more directly affected during hypoxia and its expression is regulated by mTOR activity. Additionally, we conclude that activation of PKM2 could reverse and/or rescue the deceased placental and fetal weights observed during IUGR. These results suggest that PKM2 could be a mediator of trophoblast cell invasion and its abundance influences the development of complicated pregnancies like IUGR.
Asunto(s)
Movimiento Celular/genética , Piruvato Quinasa/fisiología , Trofoblastos/fisiología , Adulto , Animales , Estudios de Casos y Controles , Adhesión Celular/genética , Células Cultivadas , Femenino , Retardo del Crecimiento Fetal/genética , Retardo del Crecimiento Fetal/metabolismo , Humanos , Recién Nacido , Isoenzimas/fisiología , Ratones , Ratones Endogámicos C57BL , Placenta/citología , Placenta/fisiología , EmbarazoRESUMEN
Th17 cell differentiation and pathogenicity depend on metabolic reprogramming inducing shifts toward glycolysis. Here, we show that the pyruvate kinase M2 (PKM2), a glycolytic enzyme required for cancer cell proliferation and tumor progression, is a key factor mediating Th17 cell differentiation and autoimmune inflammation. We found that PKM2 is highly expressed throughout the differentiation of Th17 cells in vitro and during experimental autoimmune encephalomyelitis (EAE) development. Strikingly, PKM2 is not required for the metabolic reprogramming and proliferative capacity of Th17 cells. However, T cell-specific PKM2 deletion impairs Th17 cell differentiation and ameliorates symptoms of EAE by decreasing Th17 cell-mediated inflammation and demyelination. Mechanistically, PKM2 translocates into the nucleus and interacts with STAT3, enhancing its activation and thereby increasing Th17 cell differentiation. Thus, PKM2 acts as a critical nonmetabolic regulator that fine-tunes Th17 cell differentiation and function in autoimmune-mediated inflammation.
Asunto(s)
Autoinmunidad/fisiología , Inflamación/metabolismo , Piruvato Quinasa/fisiología , Factor de Transcripción STAT3/metabolismo , Células Th17/fisiología , Animales , Diferenciación Celular , Encefalomielitis Autoinmune Experimental/metabolismo , Encefalomielitis Autoinmune Experimental/fisiopatología , Citometría de Flujo , Técnica del Anticuerpo Fluorescente , Ratones , Ratones Endogámicos C57BL , Piruvato Quinasa/metabolismo , Reacción en Cadena en Tiempo Real de la Polimerasa , Células Th17/metabolismoRESUMEN
Lymphangioleiomyomatosis (LAM) is a devastating lung disease caused by inactivating gene mutations in either TSC1 or TSC2 that result in hyperactivation of the mechanistic target of rapamycin complex 1 (mTORC1). As LAM occurs predominantly in women during their reproductive age and is exacerbated by pregnancy, the female hormonal environment, and in particular estrogen, is implicated in LAM pathogenesis and progression. However, detailed underlying molecular mechanisms are not well understood. In this study, utilizing human pulmonary LAM specimens and cell culture models of TSC2-deficient LAM patient-derived and rat uterine leiomyoma-derived cells, we tested the hypothesis that estrogen promotes the growth of mTORC1-hyperactive cells through pyruvate kinase M2 (PKM2). Estrogen increased the phosphorylation of PKM2 at Ser37 and induced the nuclear translocation of phospho-PKM2. The estrogen receptor antagonist Faslodex reversed these effects. Restoration of TSC2 inhibited the phosphorylation of PKM2 in an mTORC1 inhibitor-insensitive manner. Finally, accumulation of phosphorylated PKM2 was evident in pulmonary nodule from LAM patients. Together, our data suggest that female predominance of LAM might be at least in part attributed to estrogen stimulation of PKM2-mediated cellular metabolic alterations. Targeting metabolic regulators of PKM2 might have therapeutic benefits for women with LAM and other female-specific neoplasms.
Asunto(s)
Estrógenos/metabolismo , Piruvato Quinasa/metabolismo , Proteína 2 del Complejo de la Esclerosis Tuberosa/genética , Animales , Línea Celular Tumoral , Estrógenos/fisiología , Femenino , Humanos , Pulmón/patología , Neoplasias Pulmonares/patología , Linfangioleiomiomatosis/genética , Linfangioleiomiomatosis/fisiopatología , Diana Mecanicista del Complejo 1 de la Rapamicina , Fosforilación , Piruvato Quinasa/fisiología , Ratas , Transducción de Señal/efectos de los fármacos , Proteína 1 del Complejo de la Esclerosis Tuberosa/genética , Proteína 1 del Complejo de la Esclerosis Tuberosa/metabolismo , Proteína 2 del Complejo de la Esclerosis Tuberosa/metabolismo , Proteínas Supresoras de Tumor/genéticaRESUMEN
Pyruvate kinase M2 (PKM2) is a key glycolysis enzyme, and its effect on macrophages has not been entirely elucidated. Here, we identified that the PKM2 small-molecule agonist TEPP-46 mediated PKM2 activation by inducing the formation of PKM2 tetramer and promoted macrophage endotoxin tolerance. Lipopolysaccharide (LPS)-tolerant mice had higher expression of the PKM2 tetramer, which was associated with a reduced in vivo immune response to LPS. Pretreatment of macrophages with TEPP-46 resulted in tolerance to LPS stimulation, as demonstrated by a significant reduction in the production of TNF-α and IL-6. We found that TEPP-46 induced mitochondrial biogenesis in macrophages. Inhibition of mitochondrial biogenesis by mtTFA knockdown effectively inhibited TEPP-46-mediated macrophage tolerance to endotoxins. We discovered that TEPP-46 promoted the expression of PGC-1α and that PGC-1α was the key regulator of mitochondrial biogenesis in macrophages induced by TEPP-46. PGC-1α was negatively regulated by the PI3K/Akt signaling pathway. Knockdown of PKM2 or PGC-1α uniformly inhibited TEPP-46-mediated endotoxin tolerance by inhibiting mitochondrial biogenesis. In addition, TEPP-46 protected mice from lethal endotoxemia and sepsis. Collectively, these findings reveal novel mechanisms for the metabolic control of inflammation and for the induction of endotoxin tolerance by promoting mitochondrial biogenesis. Targeting PKM2 appears to be a new therapeutic option for the treatment of sepsis and other inflammatory diseases.
Asunto(s)
Lipopolisacáridos/farmacología , Macrófagos/efectos de los fármacos , Mitocondrias/efectos de los fármacos , Biogénesis de Organelos , Piruvato Quinasa/fisiología , Animales , Línea Celular , Macrófagos/fisiología , Ratones Endogámicos C57BL , Mitocondrias/fisiologíaRESUMEN
Bcl-2 family proteins control a decisive apoptotic event: mitochondrial outer membrane permeabilization (MOMP). To discover MOMP-regulating proteins, we expressed a library of intracellular single-chain variable fragments (scFvs) ("intrabodies") and selected for those rescuing cells from apoptosis induced by BimS (the short isoform of Bim). One anti-apoptotic intrabody, intrabody 5 (IB5), recognized pyruvate kinase M2 (PKM2), which is expressed in cancer cells. PKM2 deletion ablated this clonogenic rescue; thus, IB5 activated a latent cytoprotective function of PKM2. This resulted not from pyruvate kinase activity per se but rather from the formation of an active tetrameric conformation of PKM2. A stably tetrameric PKM2 mutant, K422R, promoted cell survival even in the absence of IB5, and IB5 further increased survival. Mitochondria isolated from IB5-expressing cells were relatively resistant to MOMP in vitro. In cells, IB5 expression up-regulated Mitofusin-1 (Mfn1) and increased mitochondrial length. Importantly, Mfn1 deficiency abrogated IB5's cytoprotective effect. PKM2's anti-apoptotic function could help explain its preferential expression in human cancer.
Asunto(s)
Permeabilidad de la Membrana Celular/fisiología , Membranas Mitocondriales/fisiología , Piruvato Quinasa/metabolismo , Secuencia de Aminoácidos , Apoptosis/fisiología , Proteína 11 Similar a Bcl2/metabolismo , Proteínas Portadoras/metabolismo , Línea Celular Tumoral , Proliferación Celular , GTP Fosfohidrolasas/metabolismo , Biblioteca de Genes , Células HEK293 , Humanos , Proteínas de la Membrana/metabolismo , Mitocondrias/metabolismo , Proteínas de Transporte de Membrana Mitocondrial/metabolismo , Isoformas de Proteínas/metabolismo , Piruvato Quinasa/fisiología , Transducción de Señal , Anticuerpos de Cadena ÚnicaRESUMEN
Pyruvate Kinase isozymes M2 (PKM2) is a glycolytic enzyme involved in glycolysis that decarboxylates phosphoenolpyruvate to pyruvate and generates ATP. PKM2 also plays a significant role in tumor growth, in cell division, angiogenesis, apoptosis and metastasis. In this study, we have investigated the role of PKM2 in cortical neurons which suffered hypoxic-ischemic encephalopathy (HIE) in newborn rats. Immunohistochemistry and Western blot analysis revealed the protein expression of PKM2 peaking at 24 h after HIE. Double immunofluorescence labeling showed that PKM2 was mainly located in the neurons of the ipsilateral cerebral cortex, not in astrocytes or microglia. The increased level of active caspase-3 and the decreased level of phosphorylated AKT (p-AKT) were consistent with the PKM2 expression. TUNEL staining assay showed that PKM2 may participate in neuronal apoptosis in the rat ipsilateral cerebral cortex. Silencing of PKM2 in primary cultures of cortical neurons using a specific siRNA reduced the expression of active caspase-3 and upregulated p-AKT expression. Taken together, the results indicate that PKM2 may be involved in neuronal apoptosis after HIE by a mechanism dependent on the inactivation of p-AKT.
Asunto(s)
Apoptosis/fisiología , Corteza Cerebral/fisiología , Hipoxia-Isquemia Encefálica/fisiopatología , Neuronas/fisiología , Piruvato Quinasa/fisiología , Animales , Animales Recién Nacidos , Encéfalo/patología , Caspasa 3/metabolismo , Corteza Cerebral/patología , Hipoxia-Isquemia Encefálica/patología , Isoenzimas/genética , Isoenzimas/fisiología , Neuronas/patología , Proteínas Proto-Oncogénicas c-akt/metabolismo , Piruvato Quinasa/genética , ARN Interferente Pequeño/genética , Ratas , Regulación hacia ArribaRESUMEN
Pyruvate Kinase M2 (PKM2) mediates metabolic reshuffling and is ubiquitously upregulated in several cancer types. The non-metabolic function of PKM2 as key nuclear kinase and modulator of gene expression is instrumental in cancer progression and tumorigenesis. Here, we attempt to discern the non-canonical function of PKM2 as an epigenetic modulator and the underlying implication of this activity. Using 5'-FAM labelled reconstituted mononucleosome we have shown that PKM2 interacts with the complex through Histone H3 and possibly obstruct the access to DNA binding factors. Subsequently, the interaction negatively impacts the ATP dependent remodeling activity of Chromodomain Helicase DNA binding protein-7 (Chd7). Chd7 remodeling activity is required to ameliorate DNA damage and is crucial to genome stability. Our study shows that PKM2 blocks the Chd7 mediated sliding of nucleosome. It can be conjectured that stalling Chd7 may lead to impaired DNA damage and increased genomic instability. We propose a mechanism in which PKM2 negatively regulate nucleosome repositioning in chromatin and may exacerbate cancer by altering the nucleosome architecture. This research is imperative to our understanding of how altered cancer metabolism can potentially modulate the gene expression and sustain incessant proliferation by tweaking the chromatin topography.
Asunto(s)
Proteínas Portadoras/fisiología , ADN Helicasas/fisiología , Proteínas de Unión al ADN/fisiología , Proteínas de la Membrana/fisiología , Nucleosomas/metabolismo , Piruvato Quinasa/fisiología , Hormonas Tiroideas/fisiología , Proteínas Portadoras/genética , Proteínas Portadoras/metabolismo , Ensamble y Desensamble de Cromatina , ADN Helicasas/genética , ADN Helicasas/metabolismo , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Regulación de la Expresión Génica , Humanos , Proteínas de la Membrana/genética , Proteínas de la Membrana/metabolismo , Nucleosomas/genética , Hormonas Tiroideas/genética , Hormonas Tiroideas/metabolismo , Proteínas de Unión a Hormona TiroideRESUMEN
Human immunodeficiency virus type 1 (HIV-1) recruits diverse cellular factors into viral particles during its morphogenesis, which apparently play roles in modulating its infectivity. In our study, proteomic techniques demonstrated that a key glycolytic protein, pyruvate kinase muscle type 2 (PKM2), is incorporated into viral particles. Here, we show that virion-packaged PKM2 significantly reduces viral infectivity by affecting the incorporation level of a cellular tRNALys3 into virions. Enhanced expression of PKM2 in HIV-1-producing cells led to a higher incorporation level of PKM2 into progeny virions without affecting the viral maturation process. Compared with the control virus, the high-level-PKM2-packaging virus showed decreased levels of both reverse transcription products and cellular tRNALys3 packaging, suggesting that the shortage of intravirion tRNALys3 suppresses reverse transcription efficiency in target cells. Interestingly, the enhanced expression of PKM2 also suppressed the virion recruitment of other nonpriming cellular tRNAs such as tRNALys1,2 and tRNAAsn, which are known to be selectively packaged into virions, without affecting the steady level of the cytoplasmic pool of those tRNAs in producer cells, suggesting that PKM2 specifically impedes the selective incorporation of tRNAs into virions. Taken together, our findings indicate that PKM2 is a vital host factor that negatively affects HIV-1 infectivity by targeting the tRNALys3-mediated initiation of reverse transcription in target cells.
Asunto(s)
VIH-1/fisiología , Piruvato Quinasa/fisiología , Células HEK293 , Humanos , Piruvato Quinasa/genética , ARN de Transferencia , Transcripción Reversa , Virión/fisiología , Ensamble de Virus , Internalización del VirusRESUMEN
Starch is the main form of energy storage in higher plants. Although several enzymes and regulators of starch biosynthesis have been defined, the complete molecular machinery remains largely unknown. Screening for irregularities in endosperm formation in rice represents valuable prospect for studying starch synthesis pathway. Here, we identified a novel rice white-core endosperm and defective grain filling mutant, ospk2, which displays significantly lower grain weight, decreased starch content and alteration of starch physicochemical properties when compared to wild-type grains. The normal starch compound granules were drastically reduced and more single granules filled the endosperm cells of ospk2. Meanwhile, the germination rate of ospk2 seeds after 1-year storage was observably reduced compared with wild-type. Map-based cloning of OsPK2 indicated that it encodes a pyruvate kinase (PK, ATP: pyruvate 2-O-phosphotransferase, EC 2.7.1.40), which catalyses an irreversible step of glycolysis. OsPK2 has a constitutive expression in rice and its protein localizes in chloroplasts. Enzyme assay showed that the protein product from expressed OsPK2 and the crude protein extracted from tissues of wild-type exhibits strong PK activity; however, the mutant presented reduced protein activity. OsPK2 (PKpα1) and three other putative rice plastidic isozymes, PKpα2, PKpß1 and PKpß2, can interact to form heteromer. Moreover, the mutation leads to multiple metabolic disorders. Altogether, these results denote new insights into the role of OsPK2 in plant seed development, especially in starch synthesis, compound granules formation and grain filling, which would be useful for genetic improvement of high yield and rice grain quality.
Asunto(s)
Grano Comestible/crecimiento & desarrollo , Endospermo/crecimiento & desarrollo , Genes de Plantas/genética , Oryza/genética , Proteínas de Plantas/genética , Piruvato Quinasa/genética , Almidón/biosíntesis , Endospermo/metabolismo , Genes de Plantas/fisiología , Oryza/enzimología , Oryza/crecimiento & desarrollo , Oryza/metabolismo , Proteínas de Plantas/metabolismo , Proteínas de Plantas/fisiología , Piruvato Quinasa/metabolismo , Piruvato Quinasa/fisiologíaRESUMEN
Pyruvate kinase muscle isozymes (PKMs) have crucial roles in regulating metabolic changes during carcinogenesis. A switch from PKM1 to PKM2 isoform was thought to lead to aerobic glycolysis promoting carcinogenesis, and was considered as one of the cancer signatures. However, recent evidence has argued against the existence of PKM isoform switch and related metabolic effects during cancer progression. We compared the effects of PKM1 and PKM2 in cell invasiveness and metastasis of pancreatic ductal adenocarcinoma (PDAC). Both PKM1 and PKM2 expression affected cell migration and invasion abilities of PDAC cells, but only knockdown of PKM2 suppressed metastasis in a xenograft model. By comparing the established PKM2 mutants in the regulation of cell invasion, we found that PKM2 may control cell mobility through its protein kinase and phopho-peptide binding abilities. Further survey for PKM2-associated proteins identified PAK2 as a possible phosphorylation target in PDAC. In vitro binding and kinase assays revealed that PKM2 directly phosphorylated PAK2 at Ser20, Ser141, and Ser192/197. Knockdown of PKM2 decreased PAK2 protein half-life by increasing ubiquitin-dependent proteasomal degradation. Moreover, we identified PAK2 as an HSP90 client protein and the mutation at Ser192/197 of PAK2 reduced PAK2-HSP90 association. Knockdown of PAK2 diminished in vitro cell mobility and in vivo metastatic ability of PKM2 overexpressed PDAC cells. PKM2 and PAK2 protein expression also positively correlated with each other in PDAC tissues. Our findings indicate that PKM2-PAK2 regulation is critical for developing metastasis in PDAC, and suggest that targeting the PKM2/HSP90/PAK2 complex has a potential therapeutic value in this deadly disease.
Asunto(s)
Carcinoma Ductal Pancreático/patología , Neoplasias Pancreáticas/patología , Piruvato Quinasa/fisiología , Quinasas p21 Activadas/metabolismo , Animales , Carcinoma Ductal Pancreático/genética , Carcinoma Ductal Pancreático/metabolismo , Movimiento Celular/genética , Células Cultivadas , Humanos , Masculino , Ratones , Ratones Endogámicos NOD , Ratones SCID , Ratones Transgénicos , Invasividad Neoplásica , Metástasis de la Neoplasia , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/metabolismo , Fosforilación/genética , Procesamiento Proteico-Postraduccional/genética , Estabilidad Proteica , Piruvato Quinasa/genética , Quinasas p21 Activadas/genéticaRESUMEN
Pyruvate kinase M2 (PKM2) has been wildly verified to modulate glycolysis in tumor cells. However, the role of PKM2 on the glycolysis of osteoarthritis (OA) chondrocytes is still unclear. In present study, we investigate the function of PKM2 on OA chondrocyte glycolysis and the collagen matrix generation in vitro. Results showed that PKM2 was upregulated in OA chondrocytes compared with healthy control chondrocytes. In OA chondrocytes, ATP expression was lower compared with healthy control chondrocytes. Loss-of-function experiment showed that PKM2 knockdown mediated by lentivirus transfection could significantly suppress the glucose consumption and lactate secretion levels and decrease glucose transporter-1 (Glut-1), lactate dehydrogenase A (LDHA), and hypoxia inducible factor 1-alpha (HIF-1α), indicating the inhibition of PKM2 knockdown on glycolysis. Moreover, Cell Counting Kit-8 (CCK-8), flow cytometry, and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) assay showed that PKM2 knockdown inhibited OA chondrocyte proliferation and promoted the apoptosis. Western blot and immunocytochemical staining showed that PKM2 knockdown downregulated the expression levels of COL2A1 and SOX-9. In summary, our results conclude that PKM2 modulates the glycolysis and extracellular matrix generation, providing the vital role of PKM2 on OA pathogenesis and a novel therapeutic target for OA.
Asunto(s)
Condrocitos/enzimología , Matriz Extracelular/enzimología , Glucólisis , Osteoartritis de la Rodilla/enzimología , Piruvato Quinasa/fisiología , Adulto , Proliferación Celular , Células Cultivadas , Inducción Enzimática , Humanos , Persona de Mediana Edad , Osteoartritis de la Rodilla/patología , Regulación hacia ArribaRESUMEN
BACKGROUNDS: Numerous studies have reported that aberrant pyruvate kinase M2 isoform (PKM2) expressed in cancer, indicating that PKM2 plays a critical role in tumor initiation and progression. Nevertheless, its prognostic value in breast cancer tumor is yet contentious. Therefore, we performed this meta-analysis to evaluate the prognostic significance of PKM2 in breast cancer. METHODS: Eligible relevant literatures were retrieved by searching PubMed, the Cochrane Library, Embase through December 2016. Articles that comparing different PKM2 expression levels in human breast cancer tissues and prognostic significance were included. Software RevMan 5.3 and STATA (Review Manager (RevMan): [Computer program]. Version 5.3. Copenhagen: The Nordic Cochrane Centre, The Cochrane Collaboration, 2014. STATA: StataCorp. 2011. Stata Statistical Software: Release 12. College Station, TX: StataCorp LP) were applied to analyze the outcomes. Pooled results were presented in hazardous ratios (HRs) of 5-year overall survival (OS), progression-free survival (PFS), and odds ratios (ORs) of clinicopathological features with 95% confidence intervals. RESULTS: Data from 6 involved studies with 895 patients were summarized. Breast cancer patients with high PKM2 had a worse OS (pooled HRâ=â1.65, 95% CIâ=â1.31-2.08, Pâ<â.001) and PFS (pooled HRâ=â2.49, 95% CIâ=â1.84-3.36, Pâ<â.00001). High PKM2 expression is related to lymph node metastasis (N1+N2+N3 vs N0, ORâ=â1.97, 95%CIâ=â1.39-2.80, Pâ=â.0001). The outcome stability was verified via sensitivity analysis. But elevated PKM2 expression was not correlated to tumor stage (T2+T3 vs T1, pooled ORâ=â0.80, 95% CIâ=â0.36-1.77, Pâ=â.58) and differential grade (G2+G3 vs G1, ORâ=â2.74, 95%CIâ=â0.76-9.84, Pâ=â.12). No publication bias was found in the included studies for OS (Begg test, Pâ=â.260; Egger test, Pâ=â.747). CONCLUSIONS: High PKM2 expression denotes worse OS and PFS in breast cancer patients, and correlate with the lymph node metastasis. However, there is no evidence for the impact of PKM2 expression on T stage and tumor differentiation.
Asunto(s)
Neoplasias de la Mama/enzimología , Neoplasias de la Mama/mortalidad , Piruvato Quinasa/fisiología , Femenino , Humanos , Pronóstico , Tasa de SupervivenciaRESUMEN
Metastasis is one of the main causes of breast cancer (BCa)-related deaths in female. It has been reported that cancer stem cell played an important role in metastasis. Here we first revealed a specific role of pyruvate kinase isozymes M2 (PKM2) in the stemness of breast cancer cells. Breast cancer tissue analysis confirmed the upregulation of PKM2 in breast cancer, and high PKM2 levels were associated with poor prognosis of breast cancer patients. Holoclone assay and colony formation assay significantly elucidated the role of PKM2 in the self-renewal of breast cancer cells. Moreover, PKM2 elevated the proportion of stem cell and the ability of sphere formation in breast cancer cells. PKM2 played its functional role in stemness by regulating ß-catenin. Collectively, we identified critical roles of PKM2 in the stemness of breast cancer cells which may elevate the therapeutic effect on breast cancer patients.
Asunto(s)
Neoplasias de la Mama/enzimología , Células Madre Neoplásicas/enzimología , Piruvato Quinasa/fisiología , Vía de Señalización Wnt , Neoplasias de la Mama/mortalidad , Neoplasias de la Mama/patología , Línea Celular Tumoral , Autorrenovación de las Células , Femenino , Expresión Génica , Humanos , Isoenzimas/fisiología , Metástasis de la Neoplasia , Pronóstico , beta Catenina/genética , beta Catenina/metabolismoRESUMEN
UNLABELLED: Hormones and their corresponding receptors are vital in controlling metabolism under normal physiologic and pathologic conditions, but less is known about their roles in the metabolism of cancer. Using a small interfering RNA screening approach, we examined the effects of silencing 20 well-known hormone receptors on the Warburg effect, specifically by measuring the production of lactate in four established hepatocellular carcinoma (HCC) cell lines. We found that silencing a variety of hormone receptors had effects on the production of this metabolite. Unexpectedly silencing of mineralocorticoid receptor (MR) significantly increased lactate production in all these HCC cell lines. Subsequent in vitro and in vivo studies showed that gain- and loss-of-function of MR significantly influenced HCC cellular proliferation, cell cycle distribution, and apoptosis. Furthermore, mechanistic studies revealed that MR as a transcriptional factor directly regulated the expression of miR-338-3p, suppressing the Warburg effects of HCC cells by targeting a key enzyme of glycolysis: pyruvate kinase, liver and red blood cells. Moreover, MR expression was significantly down-regulated in 81% of HCC patient tissues, caused by both chromosome deletion and histone deacetylation. Low expression of MR in tumor tissues was associated with poor patient prognosis. The expression level of miR-338-3p was found to positively correlate with the expression of MR in HCC tissues and to inversely correlate with expression of the enzyme pyruvate kinase, liver and red blood cells. CONCLUSION: MR affects HCC development by modulating the miR-338-3p/pyruvate kinase, liver and red blood cells axis with an ability to suppress the Warburg effect.
Asunto(s)
Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/metabolismo , Glucólisis , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/metabolismo , MicroARNs/fisiología , Piruvato Quinasa/fisiología , Receptores de Mineralocorticoides/fisiología , Carcinoma Hepatocelular/patología , Progresión de la Enfermedad , Femenino , Humanos , Neoplasias Hepáticas/patología , Masculino , Persona de Mediana Edad , Células Tumorales CultivadasRESUMEN
Cancer cells are characterized by reprogramming of energy metabolism. Over the last decade, understanding of the metabolic changes that occur in cancer has increased dramatically, with great interest in targeting metabolism for cancer therapy. Pyruvate kinase isoenzyme type M2 (abbreviations: PKM2, M2-PK) plays a key role in modulating glucose metabolism to support cell proliferation. PKM2, like other PK isoforms, catalyzes the last energy-generating step in glycolysis, but is unique in its capacity to be regulated. PKM2 is regulated at several cellular levels, including gene expression, alternative splicing and post-translational modification. In addition, PKM2 is regulated by key metabolic intermediates and interacts with more than twenty different proteins. Hence, this isoenzyme is an important regulator of glycolysis, and additionally functions in other novel roles that have recently emerged. Recent evidence indicates that intervening with the complex regulatory network of PKM2 has severe consequences on tumor cell proliferation, indicating the potential of this enzyme as a target for tumor therapy.
Asunto(s)
Neoplasias/metabolismo , Piruvato Quinasa/fisiología , Animales , Activación Enzimática , Glucólisis , Humanos , Neoplasias/tratamiento farmacológico , Procesamiento Proteico-Postraduccional , Piruvato Quinasa/antagonistas & inhibidores , Piruvato Quinasa/genéticaRESUMEN
Lysine acetylation plays an essential role in metabolism. Five individual studies have identified that a large number of cellular proteins are potentially acetylated. Notably, almost every enzyme involved in central metabolic pathways such as glycolysis, the TCA cycle, fat acid metabolism, urea cycle and glycogen metabolism, is acetylated in response to nutrition fluctuations. Metabolic reprogramming is a critical hallmark during cancer development. Tumor cells preferentially utilize glycolysis instead of oxidative phosphorylation to produce more lactate and metabolic intermediates even under normal oxygen pressure, which was first noted as the "Warburg Effect". This review focuses on recent advances in the acetylation regulation of metabolic enzymes involved in the Warburg effect, the dysfunction of acetylation regulation in tumorigenesis and their potential role in cancer metabolism therapy.
Asunto(s)
Neoplasias/metabolismo , Acetilación , Animales , Glucólisis , Humanos , Lipogénesis , Lisina/metabolismo , Piruvato Quinasa/fisiología , Especies Reactivas de Oxígeno/metabolismoRESUMEN
BACKGROUND: The early diagnosis of biliary tract cancer (BTC) remains challenging, and there are few effective therapies. This study investigated whether the M2 isotype of pyruvate kinase (M2-PK), which serves as the key regulator of cellular energy metabolism in proliferating cells, could play a role in the diagnosis and therapy of BTC. METHODS: Plasma and bile M2-PK concentrations were measured by enzyme-linked immunosorbent assay in 88 patients with BTC, 79 with benign biliary diseases, and 17 healthy controls. M2-PK expression was assayed in a BTC tissue array by immunohistochemistry. The role of M2-PK in tumor growth, invasion, and angiogenesis was evaluated in BTC cell lines by retrovirus-mediated M2-PK transfection and short hairpin RNA silencing techniques. RESULTS: Sensitivity (90.3%) and specificity (84.3%) of bile M2-PK for malignancy were significantly higher than those for plasma M2-PK and serum carbohydrate antigen 19-9. M2-PK expression was specific for cancer cells and correlated with microvessel density. M2-PK positivity was a significant independent prognostic factor by multivariable analysis. Transfection of M2-PK in a negatively expressed cell line (HuCCT-1 cells) increased cell invasion, whereas silencing in an M2-PK-positive cell line (TFK cells) decreased tumor nodule formation and cellular invasion. A significant increase in endothelial tube formation was noted when supernatants from M2-PK-transfected cells were added to an in vitro angiogenesis assay, whereas supernatants from silenced cells negated endothelial tube formation. CONCLUSIONS: Bile M2-PK is a novel tumor marker for BTC and correlates with tumor aggressiveness and poor outcome. Short hairpin RNA-mediated inhibition of M2-PK indicates the potential of M2-PK as a therapeutic target.
Asunto(s)
Neoplasias del Sistema Biliar/diagnóstico , Biomarcadores de Tumor , Carcinoma/diagnóstico , Piruvato Quinasa/fisiología , Adulto , Anciano , Anciano de 80 o más Años , Bilis/química , Bilis/metabolismo , Neoplasias del Sistema Biliar/genética , Neoplasias del Sistema Biliar/mortalidad , Neoplasias del Sistema Biliar/patología , Biomarcadores de Tumor/sangre , Biomarcadores de Tumor/genética , Biomarcadores de Tumor/metabolismo , Biomarcadores de Tumor/fisiología , Carcinoma/genética , Carcinoma/mortalidad , Carcinoma/patología , Estudios de Casos y Controles , Línea Celular Tumoral , Progresión de la Enfermedad , Femenino , Humanos , Masculino , Persona de Mediana Edad , Pronóstico , Piruvato Quinasa/sangre , Piruvato Quinasa/genética , Piruvato Quinasa/metabolismo , Análisis de SupervivenciaRESUMEN
In Clostridium thermocellum, a thermophilic anaerobic bacterium able to rapidly ferment cellulose to ethanol, pyruvate kinase (EC 2.7.1.40) is absent based on both the genome sequence and enzymatic assays. Instead, a new pathway converting phosphoenolpyruvate to pyruvate via a three-step pathway involving phosphoenolpyruvate carboxykinase, NADH-linked malate dehydrogenase, and NADP-dependent malic enzyme has been found. We examined the impact of targeted modification of enzymes associated with this pathway, termed the "malate shunt", including expression of the pyruvate kinase gene from Thermoanaerobacterium saccharolyticum, mutation of the phosphoenolpyruvate carboxykinase and deletion of malic enzyme gene. Strain YD01 with exogenous pyruvate kinase, in which phosphoenolpyruvate carboxykinase expression was diminished by modifying the start codon from ATG to GTG, exhibited 3.25-fold higher ethanol yield than the wild-type strain. A second strain, YD02 with exogenous pyruvate kinase, in which the gene for malic enzyme and part of malate dehydrogenase were deleted, had over 3-fold higher ethanol yield than the wild-type strain.
Asunto(s)
Carbono/metabolismo , Celulosa/metabolismo , Clostridium thermocellum/fisiología , Etanol/metabolismo , Mejoramiento Genético/métodos , Piruvato Quinasa/fisiología , Thermoanaerobacter/fisiología , Etanol/aislamiento & purificaciónRESUMEN
Pyruvate kinase isoform M2 (PKM2) is a glycolysis enzyme catalyzing conversion of phosphoenolpyruvate (PEP) to pyruvate by transferring a phosphate from PEP to ADP. We report here that PKM2 localizes to the cell nucleus. The levels of nuclear PKM2 correlate with cell proliferation. PKM2 activates transcription of MEK5 by phosphorylating stat3 at Y705. In vitro phosphorylation assays show that PKM2 is a protein kinase using PEP as a phosphate donor. ADP competes with the protein substrate binding, indicating that the substrate may bind to the ADP site of PKM2. Our experiments suggest that PKM2 dimer is an active protein kinase, while the tetramer is an active pyruvate kinase. Expression of a PKM2 mutant that exists as a dimer promotes cell proliferation, indicating that protein kinase activity of PKM2 plays a role in promoting cell proliferation. Our study reveals an important link between metabolism alteration and gene expression during tumor transformation and progression.