RESUMO
Ewing sarcoma is characterized by a pathognomonic chromosomal translocation that generates the EWSR1-FLI1 chimeric transcription factor. The transcriptional targets of EWSR1-FLI1 that are essential for tumorigenicity are incompletely defined. Here, we found that EWSR1-FLI1 modulates the expression of cancer/testis (CT) antigen genes, whose expression is biased to the testes but is also activated in cancer. Among these CT antigens, fetal and adult testis expressed 1 (FATE1) is most robustly induced. EWSR1-FLI1 associates with the GGAA repeats in the proximal promoter of FATE1, which exhibits accessible chromatin exclusively in mesenchymal progenitor cells (MPCs) and Ewing sarcoma cells. Expression of EWSR1-FLI1 in non-Ewing sarcoma cells and in MPCs enhances FATE1 mRNA and protein expression. Conversely, depletion of EWSR1-FLI1 in Ewing sarcoma cells leads to a loss of FATE1 expression. Importantly, we found that FATE1 is required for survival and anchorage-independent growth in Ewing sarcoma cells via attenuating the accumulation of BNIP3L, a BH3-only protein that is toxic when stabilized. This action appears to be mediated by the E3 ligase RNF183. We propose that engaging FATE1 function can permit the bypass of cell death mechanisms that would otherwise inhibit tumor progression.
Assuntos
Neoplasias Ósseas/genética , Proteínas de Ligação a DNA/genética , Proteínas de Fusão Oncogênica/genética , Sarcoma de Ewing/genética , Fatores de Transcrição/genética , Neoplasias Ósseas/metabolismo , Linhagem Celular Tumoral , Sobrevivência Celular , Proteínas de Ligação a DNA/metabolismo , Regulação Neoplásica da Expressão Gênica , Células HeLa , Células Hep G2 , Humanos , Proteínas de Membrana/metabolismo , Regiões Promotoras Genéticas , Proteínas Proto-Oncogênicas/metabolismo , Sarcoma de Ewing/metabolismo , Fatores de Transcrição/metabolismo , Proteínas Supressoras de Tumor/metabolismo , Ubiquitina-Proteína Ligases/metabolismoRESUMO
Tumours frequently activate genes whose expression is otherwise biased to the testis, collectively known as cancer-testis antigens (CTAs). The extent to which CTA expression represents epiphenomena or confers tumorigenic traits is unknown. In this study, to address this, we implemented a multidimensional functional genomics approach that incorporates 7 different phenotypic assays in 11 distinct disease settings. We identify 26 CTAs that are essential for tumor cell viability and/or are pathological drivers of HIF, WNT or TGFß signalling. In particular, we discover that Foetal and Adult Testis Expressed 1 (FATE1) is a key survival factor in multiple oncogenic backgrounds. FATE1 prevents the accumulation of the stress-sensing BH3-only protein, BCL-2-Interacting Killer (BIK), thereby permitting viability in the presence of toxic stimuli. Furthermore, ZNF165 promotes TGFß signalling by directly suppressing the expression of negative feedback regulatory pathways. This action is essential for the survival of triple negative breast cancer cells in vitro and in vivo. Thus, CTAs make significant direct contributions to tumour biology.
Assuntos
Antígenos de Neoplasias/genética , Carcinogênese/genética , Proteínas de Ligação a DNA/genética , Regulação Neoplásica da Expressão Gênica , Neoplasias/genética , Fatores de Transcrição/genética , Fator de Crescimento Transformador beta/genética , Adenocarcinoma/genética , Adenocarcinoma/imunologia , Adenocarcinoma/mortalidade , Adenocarcinoma de Pulmão , Animais , Proteínas Reguladoras de Apoptose/genética , Linhagem Celular , Linhagem Celular Tumoral , Sobrevivência Celular , Neoplasias Colorretais/genética , Neoplasias Colorretais/imunologia , Neoplasias Colorretais/mortalidade , Imunofluorescência , Células HCT116 , Células HEK293 , Humanos , Immunoblotting , Técnicas In Vitro , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/imunologia , Neoplasias Pulmonares/mortalidade , Proteínas de Membrana/genética , Camundongos Endogâmicos NOD , Proteínas Mitocondriais , Transplante de Neoplasias , Neoplasias/imunologia , Neoplasias/mortalidade , Prognóstico , Modelos de Riscos Proporcionais , Reação em Cadeia da Polimerase em Tempo Real , Transdução de Sinais , Proteína Smad7/genética , Fator de Crescimento Transformador beta/imunologia , Neoplasias de Mama Triplo Negativas/genética , Neoplasias de Mama Triplo Negativas/imunologia , Neoplasias de Mama Triplo Negativas/mortalidade , Ubiquitina-Proteína Ligases/genética , Via de Sinalização WntRESUMO
Non-small cell lung cancer (NSCLC) is notorious for its paltry responses to first-line therapeutic regimens. In contrast to acquired chemoresistance, little is known about the molecular underpinnings of the intrinsic resistance of chemo-naïve NSCLC. Here we report that intrinsic resistance to paclitaxel in NSCLC occurs at a cell-autonomous level because of the uncoupling of mitotic defects from apoptosis. To identify components that permit escape from mitotic stress-induced death, we used a genome-wide RNAi-based strategy, which combines a high-throughput toxicity screen with a live-cell imaging platform to measure mitotic fate. This strategy revealed that prolonging mitotic arrest with a small molecule inhibitor of the APC/cyclosome could sensitize otherwise paclitaxel-resistant NSCLC. We also defined novel roles for CASC1 and TRIM69 in supporting resistance to spindle poisons. CASC1, which is frequently co-amplified with KRAS in lung tumors, is essential for microtubule polymerization and satisfaction of the spindle assembly checkpoint. TRIM69, which associates with spindle poles and promotes centrosomal clustering, is essential for formation of a bipolar spindle. Notably, RNAi-mediated attenuation of CASC1 or TRIM69 was sufficient to inhibit tumor growth in vivo. On the basis of our results, we hypothesize that tumor evolution selects for a permissive mitotic checkpoint, which may promote survival despite chromosome segregation errors. Attacking this adaptation may restore the apoptotic consequences of mitotic damage to permit the therapeutic eradication of drug-resistant cancer cells.
Assuntos
Carcinoma Pulmonar de Células não Pequenas/metabolismo , Neoplasias Pulmonares/metabolismo , Mitose , Estresse Fisiológico , Animais , Antineoplásicos/farmacologia , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/patologia , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Pontos de Checagem do Ciclo Celular/genética , Morte Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Modelos Animais de Doenças , Resistencia a Medicamentos Antineoplásicos/genética , Feminino , Xenoenxertos , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Camundongos , Microtúbulos/metabolismo , Mitose/efeitos dos fármacos , Mitose/genética , Paclitaxel/farmacologia , Ligação Proteica , Interferência de RNA , Fuso Acromático/metabolismo , Estresse Fisiológico/efeitos dos fármacos , Estresse Fisiológico/genética , Proteínas com Motivo Tripartido , Carga Tumoral/efeitos dos fármacos , Ubiquitina-Proteína Ligases/genética , Ubiquitina-Proteína Ligases/metabolismoRESUMO
While the expression of genes that are normally involved in spermatogenesis is frequently detected in tumors, the extent to which these gene products are required for neoplastic behaviors is unclear. To begin to address their functional relevance to tumorigenesis, we identified a cohort of proteins which display synthetic lethality with paclitaxel in non-small-cell lung cancer and whose expression is biased toward testes and tumors. Remarkably, these testis proteins, FMR1NB, NXF2, MAGEA5, FSIP1, and STARD6, are required for accurate chromosome segregation in tumor cells. Their individual depletion enhances the generation of multipolar spindles, increases mitotic transit time, and induces micronucleation in response to an otherwise innocuous dose of paclitaxel. The underlying basis for abnormal mitosis is an alteration in microtubule function, as their depletion increases microtubule cytaster formation and disrupts microtubule stability. Given these observations, we hypothesize that reactivated testis proteins may represent unique tumor cell vulnerabilities which, if targeted, could enhance responsiveness to antimitotic therapy. Indeed, we demonstrate that combining paclitaxel with a small-molecule inhibitor of the gametogenic and tumor cell mitotic protein TACC3 leads to enhanced centrosomal abnormalities, activation of death programs, and loss of anchorage-independent growth.
