RESUMO
Accumulating evidence indicates that the MDM2 oncoprotein promotes tumorigenesis beyond its canonical negative effects on the p53 tumor suppressor, but these p53-independent functions remain poorly understood. Here, we show that a fraction of endogenous MDM2 is actively imported in mitochondria to control respiration and mitochondrial dynamics independently of p53. Mitochondrial MDM2 represses the transcription of NADH-dehydrogenase 6 (MT-ND6) in vitro and in vivo, impinging on respiratory complex I activity and enhancing mitochondrial ROS production. Recruitment of MDM2 to mitochondria increases during oxidative stress and hypoxia. Accordingly, mice lacking MDM2 in skeletal muscles exhibit higher MT-ND6 levels, enhanced complex I activity, and increased muscular endurance in mild hypoxic conditions. Furthermore, increased mitochondrial MDM2 levels enhance the migratory and invasive properties of cancer cells. Collectively, these data uncover a previously unsuspected function of the MDM2 oncoprotein in mitochondria that play critical roles in skeletal muscle physiology and may contribute to tumor progression.
Assuntos
Carcinoma Pulmonar de Células não Pequenas/patologia , Transformação Celular Neoplásica/patologia , Complexo I de Transporte de Elétrons/metabolismo , Regulação Neoplásica da Expressão Gênica , Mitocôndrias/patologia , Proteínas Proto-Oncogênicas c-mdm2/metabolismo , Proteína Supressora de Tumor p53/metabolismo , Adenocarcinoma/genética , Adenocarcinoma/metabolismo , Adenocarcinoma/patologia , Animais , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Movimento Celular , Transformação Celular Neoplásica/genética , Transformação Celular Neoplásica/metabolismo , Complexo I de Transporte de Elétrons/genética , Genoma Mitocondrial , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Mitocôndrias/genética , Mitocôndrias/metabolismo , Invasividade Neoplásica , Estresse Oxidativo , Proteínas Proto-Oncogênicas c-mdm2/genética , Transdução de Sinais , Transcrição Gênica , Células Tumorais Cultivadas , Proteína Supressora de Tumor p53/genética , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Well-differentiated and dedifferentiated liposarcomas (LPSs) are characterized by a systematic amplification of the MDM2 oncogene, which encodes a key negative regulator of the p53 pathway. The molecular mechanisms underlying MDM2 overexpression while sparing wild-type p53 in LPS remain poorly understood. Here, we show that the p53-independent metabolic functions of chromatin-bound MDM2 are exacerbated in LPS and mediate an addiction to serine metabolism that sustains nucleotide synthesis and tumor growth. Treatment of LPS cells with Nutlin-3A, a pharmacological inhibitor of the MDM2-p53 interaction, stabilized p53 but unexpectedly enhanced MDM2-mediated control of serine metabolism by increasing its recruitment to chromatin, likely explaining the poor clinical efficacy of this class of MDM2 inhibitors. In contrast, genetic or pharmacological inhibition of chromatin-bound MDM2 by SP141, a distinct MDM2 inhibitor triggering its degradation, or interfering with de novo serine synthesis, impaired LPS growth both in vitro and in clinically relevant patient-derived xenograft models. Our data indicate that targeting MDM2 functions in serine metabolism represents a potential therapeutic strategy for LPS.
Assuntos
Antineoplásicos , Lipossarcoma , Antineoplásicos/uso terapêutico , Humanos , Lipossarcoma/tratamento farmacológico , Lipossarcoma/genética , Proteínas Proto-Oncogênicas c-mdm2/metabolismo , Serina/uso terapêutico , Proteína Supressora de Tumor p53/genéticaRESUMO
Resistance to 4-hydroxy-tamoxifen (OHT), which appears in breast cancer cells after long-term antiestrogen treatment, may involve irreversible changes of gene expression. We previously developed a MCF-7 derived cell line (MVLN), in which OHT rapidly and irreversibly inactivates the expression of an estrogen-regulated luciferase transgene (Vit-tk-luciferase). In chromatin immunoprecipitation experiments, heterochromatin protein 1 (HP1alpha) was found to be associated with the Vit-tk-luciferase transgene, only when it was inactivated by OHT treatment. Chimeras composed of either HP1alpha or the Krupple-associated box (KRAB) module of KOX-1 protein (known to repress gene expression by recruitment of HP1 proteins), fused to the estrogen receptor (ER)-DNA binding domain (DBD) and the androgen receptor (AR)-ligand binding domain (LBD) were generated and appeared as potent transcriptional repressors. In stably transfected MVLN cells, irreversible inactivation of the luciferase transgene expression obtained with HP1alpha-ER(DBD)-AR(LBD) was partial, whereas inactivation obtained with KRAB-ER(DBD)-AR(LBD) was comparable to that obtained with OHT, although with a slower kinetics. Altogether, these data suggest that HP1alpha is involved in the silencing effects associated with long-term OHT treatments.
Assuntos
Neoplasias da Mama/genética , Proteínas Cromossômicas não Histona/metabolismo , Antagonistas de Estrogênios/farmacologia , Inativação Gênica , Tamoxifeno/análogos & derivados , Transcrição Gênica/efeitos dos fármacos , Neoplasias da Mama/imunologia , Neoplasias da Mama/metabolismo , Imunoprecipitação da Cromatina , Homólogo 5 da Proteína Cromobox , Proteínas de Ligação a DNA/análise , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Resistencia a Medicamentos Antineoplásicos/genética , Humanos , Fatores de Transcrição Kruppel-Like , Luciferases/análise , Luciferases/genética , Isoformas de Proteínas/metabolismo , Receptores Androgênicos/análise , Receptores Androgênicos/genética , Receptores Androgênicos/metabolismo , Receptores de Estrogênio/análise , Receptores de Estrogênio/genética , Receptores de Estrogênio/metabolismo , Proteínas Recombinantes de Fusão/análise , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Proteínas Repressoras/análise , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Tamoxifeno/farmacologia , Transgenes , Células Tumorais CultivadasRESUMO
Receptor interacting protein 140 (RIP140) is a negative transcriptional regulator of nuclear hormone receptors that is required for the maintenance of energy homeostasis and ovulation. In this study, we investigated the mechanisms by which RIP140 expression is controlled by estrogens in breast cancer cells. We first analyzed by real time reverse transcription-polymerase chain reaction the regulation of RIP140 mRNA accumulation by estrogen receptor (ER) ligands in MCF-7 cells. We showed that the induction by estradiol (E2) was rapid and did not affect the apparent stability of the mRNA, suggesting a direct transcriptional regulation. To further study the underlying regulatory mechanisms, we then characterized the human RIP140 gene. We identified several noncoding exons with alternative splicing and localized the promoter region more than 100 kilobases upstream from the coding exon. Although we mapped a perfect consensus estrogen response element able to bind ERalpha in gel shift and in chromatin immunoprecipitation experiments, the effect of E2 on RIP140 gene transcription was very modest. This might result at least in part from the presence of an overlapping aryl hydrocarbon receptor (AhR) binding site, which interfered with the E2 response on both the transiently transfected reporter construct and the accumulation of the endogenous RIP140 mRNA. Altogether, our data indicate that the RIP140 gene exhibits a complex structure with several noncoding exons and supports transcriptional cross-talk and feedback involving the ERalpha and AhR nuclear receptors.