RESUMEN
Deregulated expression of glycolytic enzymes contributes not only to the increased energy demands of transformed cells but also has non-glycolytic roles in tumors. However, the contribution of these non-glycolytic functions in tumor progression remains poorly defined. Here, we show that elevated expression of glyceraldehyde-3-phosphate dehydrogenase (GAPDH), but not of other glycolytic enzymes tested, increased aggressiveness and vascularization of non-Hodgkin's lymphoma. Elevated GAPDH expression was found to promote nuclear factor-κB (NF-κB) activation via binding to tumor necrosis factor receptor-associated factor-2 (TRAF2), enhancing the transcription and the activity of hypoxia-inducing factor-1α (HIF-1α). Consistent with this, inactive mutants of GAPDH failed to bind TRAF2, enhance HIF-1 activity or promote lymphomagenesis. Furthermore, elevated expression of gapdh mRNA in biopsies from diffuse large B-cell non-Hodgkin's lymphoma patients correlated with high levels of hif-1α, vegf-a, nfkbia mRNA and CD31 staining. Collectively, these data indicate that deregulated GAPDH expression promotes NF-κB-dependent induction of HIF-1α and has a key role in lymphoma vascularization and aggressiveness.
Asunto(s)
Regulación Enzimológica de la Expresión Génica , Regulación Neoplásica de la Expresión Génica , Gliceraldehído 3-Fosfato Deshidrogenasa (NADP+)/metabolismo , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Linfoma no Hodgkin/metabolismo , Subunidad p50 de NF-kappa B/metabolismo , Animales , Biopsia , Línea Celular Tumoral , Inhibidores Enzimáticos/química , Células HeLa , Humanos , Linfoma/metabolismo , Ratones , Ratones Transgénicos , Fenotipo , Proteínas Proto-Oncogénicas c-myc/metabolismo , Factor 2 Asociado a Receptor de TNF/metabolismo , Factor A de Crecimiento Endotelial Vascular/metabolismoRESUMEN
Increased glucose catabolism and resistance to cell death are hallmarks of cancers, but the link between them remains elusive. Remarkably, under conditions where caspases are inhibited, the process of cell death is delayed but rarely blocked, leading to the occurrence of caspase-independent cell death (CICD). Escape from CICD is particularly relevant in the context of cancer as apoptosis inhibition only is often not sufficient to allow oncogenic transformation. While most glycolytic enzymes are overexpressed in tumors, glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is of particular interest as it can allow cells to recover from CICD. Here, we show that GAPDH, but no other glycolytic enzymes tested, when overexpressed could bind to active Akt and limit its dephosphorylation. Active Akt prevents FoxO nuclear localization, which precludes Bcl-6 expression and leads to Bcl-xL overexpression. The GAPDH-dependent Bcl-xL overexpression is able to protect a subset of mitochondria from permeabilization that are required for cellular survival from CICD. Thus, our work suggests that GAPDH overexpression could induce Bcl-xL overexpression and protect cells from CICD-induced chemotherapy through preservation of intact mitochondria that may facilitate tumor survival and chemotherapeutic resistance.
Asunto(s)
Apoptosis/fisiología , Caspasas/fisiología , Gliceraldehído 3-Fosfato Deshidrogenasa (NADP+)/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Regulación hacia Arriba/fisiología , Proteína bcl-X/metabolismo , Muerte Celular/fisiología , Línea Celular Tumoral , Supervivencia Celular/fisiología , Células HEK293 , Células HeLa , Humanos , Mitocondrias/fisiología , Fosfoglicerato Quinasa/fisiología , Fosfopiruvato Hidratasa/fisiología , Unión Proteica/fisiologíaAsunto(s)
Apoptosis/efectos de los fármacos , Compuestos de Bifenilo/farmacología , Linfoma/patología , Nitrofenoles/farmacología , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Sulfonamidas/farmacología , Línea Celular Tumoral , Resistencia a Antineoplásicos , Glucólisis , Humanos , Proteína 1 de la Secuencia de Leucemia de Células Mieloides , Piperazinas/farmacologíaRESUMEN
Most cancer cells exhibit increased glycolysis for generation of their energy supply. This specificity could be used to preferentially kill these cells. In this study, we identified the signaling pathway initiated by glycolysis inhibition that results in sensitization to death receptor (DR)-induced apoptosis. We showed, in several human cancer cell lines (such as Jurkat, HeLa, U937), that glucose removal or the use of nonmetabolizable form of glucose (2-deoxyglucose) dramatically enhances apoptosis induced by Fas or by tumor necrosis factor-related apoptosis-inducing ligand. This sensitization is controlled through the adenosine monophosphate (AMP)-activated protein kinase (AMPK), which is the central energy-sensing system of the cell. We established the fact that AMPK is activated upon glycolysis block resulting in mammalian target of rapamycin (mTOR) inhibition leading to Mcl-1 decrease, but no other Bcl-2 anti-apoptotic members. Interestingly, we determined that, upon glycolysis inhibition, the AMPK-mTOR pathway controlled Mcl-1 levels neither through transcriptional nor through posttranslational mechanism but rather by controlling its translation. Therefore, our results show a novel mechanism for the sensitization to DR-induced apoptosis linking glucose metabolism to Mcl-1 downexpression. In addition, this study provides a rationale for the combined use of DR ligands with AMPK activators or mTOR inhibitors in the treatment of human cancers.
Asunto(s)
Proteínas Quinasas Activadas por AMP/metabolismo , Apoptosis/fisiología , Glucólisis/fisiología , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Receptores de Muerte Celular/metabolismo , Proteínas Quinasas Activadas por AMP/antagonistas & inhibidores , Aminoimidazol Carboxamida/análogos & derivados , Aminoimidazol Carboxamida/farmacología , Anticuerpos/inmunología , Anticuerpos/farmacología , Apoptosis/efectos de los fármacos , Western Blotting , Desoxiglucosa/farmacología , Activación Enzimática/efectos de los fármacos , Glucosa/farmacología , Glucólisis/efectos de los fármacos , Células HeLa , Humanos , Péptidos y Proteínas de Señalización Intracelular/antagonistas & inhibidores , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Células Jurkat , Modelos Biológicos , Proteína 1 de la Secuencia de Leucemia de Células Mieloides , Biosíntesis de Proteínas/efectos de los fármacos , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Proto-Oncogénicas c-bcl-2/genética , Pirazoles/farmacología , Pirimidinas/farmacología , Interferencia de ARN , Receptores de Muerte Celular/inmunología , Proteínas Recombinantes/farmacología , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Ribonucleótidos/farmacología , Sirolimus/farmacología , Ligando Inductor de Apoptosis Relacionado con TNF/genética , Ligando Inductor de Apoptosis Relacionado con TNF/metabolismo , Ligando Inductor de Apoptosis Relacionado con TNF/farmacología , Serina-Treonina Quinasas TOR , Células U937 , Receptor fas/inmunología , Receptor fas/metabolismoRESUMEN
In three different endothelial cell (EC) cultures (primary human umbilical cord vein, so-called HUVEC; and immortalized cell lines HBMEC and EA-hy-926), the effects of different xenobiotics were studied in order to standardize vascular EC models for in vitro pharmacotoxicological studies. Cell characteristics were first investigated by the production and the mRNA levels of known endothelial markers in the three EC culture models. EC secretory products, tissue plasminogen activator (tPA) and von Willebrand factor (vWF), were present in the supernatant of the immortalized cell lines. The mRNA levels of vWF, tPA, platelet endothelial cell adhesion molecule-1 (PECAM-1/CD31), and beta -integrin subunit, which are involved in the control of platelet function, coagulation, and fibrinolysis as well as in cell-matrix interactions, were investigated in all EC types. For at least three parameters, cultured cells provided marked characteristics of EC phenotype, in HUVEC and in immortalized cell lines, regardless of their origin from the macro- or microcirculation. Toxicity experiments were assessed after 24 h exposure to cadmium, cyclosporin A and cisplatin by MTT assay. These experiments show nonsignificant difference in susceptibility to cyclosporin A and cadmium on HUVEC, HBMEC, and EA-hy-926. However, HBMEC, seems to be highly susceptible to cisplatin compared to HUVEC, the latter being more sensitive than EA-hy-926. For experiments conducted with cyclosporin and cadmium, cell lines could constitute an alternative material for routine cytotoxicity studies.