RESUMEN
Intracellular iron levels are strictly regulated to support homeostasis and avoid iron-mediated ROS production. Loss of iron-sulfur cluster (ISC) synthesis can increase iron loading and promote cell death by ferroptosis. Iron-responsive element-binding proteins IRP1 and IRP2 posttranscriptionally regulate iron homeostasis. IRP1 binding to target mRNAs is competitively regulated by ISC occupancy. However, IRP2 is principally thought to be regulated at the protein level via E3 ubiquitin ligase FBXL5-mediated degradation. Here, we show that ISC synthesis suppression can activate IRP2 and promote ferroptosis sensitivity via a previously unidentified mechanism. At tissue-level O2 concentrations, ISC deficiency enhances IRP2 binding to target mRNAs independent of IRP1, FBXL5, and changes in IRP2 protein level. Deletion of both IRP1 and IRP2 abolishes the iron-starvation response, preventing its activation by ISC synthesis inhibition. These findings will inform strategies to manipulate ferroptosis sensitivity and help illuminate the mechanism underlying ISC biosynthesis disorders, such as Friedreich's ataxia.
RESUMEN
Knowledge of fundamental differences between breast cancer subtypes has driven therapeutic advances; however, basal-like breast cancer (BLBC) remains clinically intractable. Because BLBC exhibits alterations in DNA repair enzymes and cell-cycle checkpoints, elucidation of factors enabling the genomic instability present in this subtype has the potential to reveal novel anti-cancer strategies. Here, we demonstrate that BLBC is especially sensitive to suppression of iron-sulfur cluster (ISC) biosynthesis and identify DNA polymerase epsilon (POLE) as an ISC-containing protein that underlies this phenotype. In BLBC cells, POLE suppression leads to replication fork stalling, DNA damage, and a senescence-like state or cell death. In contrast, luminal breast cancer and non-transformed mammary cells maintain viability upon POLE suppression but become dependent upon an ATR/CHK1/CDC25A/CDK2 DNA damage response axis. We find that CDK1/2 targets exhibit hyperphosphorylation selectively in BLBC tumors, indicating that CDK2 hyperactivity is a genome integrity vulnerability exploitable by targeting POLE.
Asunto(s)
Neoplasias de la Mama/patología , Carcinoma Basocelular/patología , Quinasa 2 Dependiente de la Ciclina/metabolismo , ADN Polimerasa II/metabolismo , Inestabilidad Genómica , Proteínas de Unión a Poli-ADP-Ribosa/metabolismo , Animales , Apoptosis , Neoplasias de la Mama/genética , Neoplasias de la Mama/metabolismo , Carcinoma Basocelular/genética , Carcinoma Basocelular/metabolismo , Ciclo Celular , Proliferación Celular , Quinasa 2 Dependiente de la Ciclina/genética , Daño del ADN , ADN Polimerasa II/genética , Femenino , Humanos , Ratones , Ratones Endogámicos NOD , Fosforilación , Proteínas de Unión a Poli-ADP-Ribosa/genética , Transducción de Señal , Células Tumorales CultivadasRESUMEN
Upon glucose restriction, eukaryotic cells upregulate oxidative metabolism to maintain homeostasis. Using genetic screens, we find that the mitochondrial serine hydroxymethyltransferase (SHMT2) is required for robust mitochondrial oxygen consumption and low glucose proliferation. SHMT2 catalyzes the first step in mitochondrial one-carbon metabolism, which, particularly in proliferating cells, produces tetrahydrofolate (THF)-conjugated one-carbon units used in cytoplasmic reactions despite the presence of a parallel cytoplasmic pathway. Impairing cytoplasmic one-carbon metabolism or blocking efflux of one-carbon units from mitochondria does not phenocopy SHMT2 loss, indicating that a mitochondrial THF cofactor is responsible for the observed phenotype. The enzyme MTFMT utilizes one such cofactor, 10-formyl THF, producing formylmethionyl-tRNAs, specialized initiator tRNAs necessary for proper translation of mitochondrially encoded proteins. Accordingly, SHMT2 null cells specifically fail to maintain formylmethionyl-tRNA pools and mitochondrially encoded proteins, phenotypes similar to those observed in MTFMT-deficient patients. These findings provide a rationale for maintaining a compartmentalized one-carbon pathway in mitochondria.
Asunto(s)
Neoplasias de la Mama/genética , Neoplasias de la Mama/patología , Glicina Hidroximetiltransferasa/metabolismo , Mitocondrias/genética , Iniciación de la Cadena Peptídica Traduccional , ARN de Transferencia de Metionina/química , Serina/química , Animales , Apoptosis , Neoplasias de la Mama/metabolismo , Sistemas CRISPR-Cas , Proliferación Celular , Citosol/metabolismo , Femenino , Glicina Hidroximetiltransferasa/antagonistas & inhibidores , Glicina Hidroximetiltransferasa/genética , Humanos , Ratones , Ratones Endogámicos NOD , Ratones SCID , Mitocondrias/efectos de los fármacos , Mitocondrias/metabolismo , Procesamiento Proteico-Postraduccional , ARN de Transferencia de Metionina/genética , ARN de Transferencia de Metionina/metabolismo , Serina/genética , Serina/metabolismo , Tetrahidrofolatos/farmacología , Células Tumorales Cultivadas , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Environmental nutrient levels impact cancer cell metabolism, resulting in context-dependent gene essentiality. Here, using loss-of-function screening based on RNA interference, we show that environmental oxygen levels are a major driver of differential essentiality between in vitro model systems and in vivo tumours. Above the 3-8% oxygen concentration typical of most tissues, we find that cancer cells depend on high levels of the iron-sulfur cluster biosynthetic enzyme NFS1. Mammary or subcutaneous tumours grow despite suppression of NFS1, whereas metastatic or primary lung tumours do not. Consistent with a role in surviving the high oxygen environment of incipient lung tumours, NFS1 lies in a region of genomic amplification present in lung adenocarcinoma and is most highly expressed in well-differentiated adenocarcinomas. NFS1 activity is particularly important for maintaining the iron-sulfur co-factors present in multiple cell-essential proteins upon exposure to oxygen compared to other forms of oxidative damage. Furthermore, insufficient iron-sulfur cluster maintenance robustly activates the iron-starvation response and, in combination with inhibition of glutathione biosynthesis, triggers ferroptosis, a non-apoptotic form of cell death. Suppression of NFS1 cooperates with inhibition of cysteine transport to trigger ferroptosis in vitro and slow tumour growth. Therefore, lung adenocarcinomas select for expression of a pathway that confers resistance to high oxygen tension and protects cells from undergoing ferroptosis in response to oxidative damage.
Asunto(s)
Liasas de Carbono-Azufre/metabolismo , Muerte Celular , Proteínas Hierro-Azufre/metabolismo , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patología , Animales , Liasas de Carbono-Azufre/genética , Carcinoma de Pulmón de Células no Pequeñas/genética , Carcinoma de Pulmón de Células no Pequeñas/metabolismo , Carcinoma de Pulmón de Células no Pequeñas/patología , Muerte Celular/genética , Línea Celular Tumoral , Cisteína/metabolismo , Glutatión/biosíntesis , Humanos , Neoplasias Pulmonares/genética , Ratones , Metástasis de la Neoplasia/genética , Metástasis de la Neoplasia/patología , Estrés Oxidativo/efectos de los fármacos , Oxígeno/metabolismo , Oxígeno/farmacología , Interferencia de ARNRESUMEN
During tumorigenesis, the high metabolic demand of cancer cells results in increased production of reactive oxygen species. To maintain oxidative homeostasis, tumor cells increase their antioxidant production through hyperactivation of the NRF2 pathway, which promotes tumor cell growth. Despite the extensive characterization of NRF2-driven metabolic rewiring, little is known about the metabolic liabilities generated by this reprogramming. Here, we show that activation of NRF2, in either mouse or human cancer cells, leads to increased dependency on exogenous glutamine through increased consumption of glutamate for glutathione synthesis and glutamate secretion by xc- antiporter system. Together, this limits glutamate availability for the tricarboxylic acid cycle and other biosynthetic reactions creating a metabolic bottleneck. Cancers with genetic or pharmacological activation of the NRF2 antioxidant pathway have a metabolic imbalance between supporting increased antioxidant capacity over central carbon metabolism, which can be therapeutically exploited.