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1.
Genes Dev ; 38(7-8): 336-353, 2024 May 21.
Artigo em Inglês | MEDLINE | ID: mdl-38744503

RESUMO

High levels of H2A.Z promote melanoma cell proliferation and correlate with poor prognosis. However, the role of the two distinct H2A.Z histone chaperone complexes SRCAP and P400-TIP60 in melanoma remains unclear. Here, we show that individual subunit depletion of SRCAP, P400, and VPS72 (YL1) results in not only the loss of H2A.Z deposition into chromatin but also a reduction of H4 acetylation in melanoma cells. This loss of H4 acetylation is particularly found at the promoters of cell cycle genes directly bound by H2A.Z and its chaperones, suggesting a coordinated regulation between H2A.Z deposition and H4 acetylation to promote their expression. Knockdown of each of the three subunits downregulates E2F1 and its targets, resulting in a cell cycle arrest akin to H2A.Z depletion. However, unlike H2A.Z deficiency, loss of the shared H2A.Z chaperone subunit YL1 induces apoptosis. Furthermore, YL1 is overexpressed in melanoma tissues, and its upregulation is associated with poor patient outcome. Together, these findings provide a rationale for future targeting of H2A.Z chaperones as an epigenetic strategy for melanoma treatment.


Assuntos
Proliferação de Células , Regulação Neoplásica da Expressão Gênica , Histonas , Melanoma , Humanos , Melanoma/genética , Proliferação de Células/genética , Linhagem Celular Tumoral , Histonas/metabolismo , Histonas/genética , Acetilação , Apoptose/genética , Fator de Transcrição E2F1/metabolismo , Fator de Transcrição E2F1/genética , Chaperonas Moleculares/metabolismo , Chaperonas Moleculares/genética
2.
bioRxiv ; 2023 Nov 27.
Artigo em Inglês | MEDLINE | ID: mdl-38076914

RESUMO

High levels of H2A.Z promote melanoma cell proliferation and correlate with poor prognosis. However, the role of the two distinct H2A.Z histone chaperone complexes, SRCAP and P400-TIP60, in melanoma remains unclear. Here, we show that individual depletion of SRCAP, P400, and VPS72 (YL1) not only results in loss of H2A.Z deposition into chromatin, but also a striking reduction of H4 acetylation in melanoma cells. This loss of H4 acetylation is found at the promoters of cell cycle genes directly bound by H2A.Z and its chaperones, suggesting a highly coordinated regulation between H2A.Z deposition and H4 acetylation to promote their expression. Knockdown of each of the three subunits downregulates E2F1 and its targets, resulting in a cell cycle arrest akin to H2A.Z depletion. However, unlike H2A.Z deficiency, loss of the shared H2A.Z chaperone subunit YL1 induces apoptosis. Furthermore, YL1 is overexpressed in melanoma tissues, and its upregulation is associated with poor patient outcome. Together, these findings provide a rationale for future targeting of H2A.Z chaperones as an epigenetic strategy for melanoma treatment.

3.
EMBO Mol Med ; 15(12): e17719, 2023 Dec 07.
Artigo em Inglês | MEDLINE | ID: mdl-37966164

RESUMO

Metastatic uveal melanomas are highly resistant to all existing treatments. To address this critical issue, we performed a kinome-wide CRISPR-Cas9 knockout screen, which revealed the LKB1-SIK2 module in restraining uveal melanoma tumorigenesis. Functionally, LKB1 loss enhances proliferation and survival through SIK2 inhibition and upregulation of the sodium/calcium (Na+ /Ca2+ ) exchanger SLC8A1. This signaling cascade promotes increased levels of intracellular calcium and mitochondrial reactive oxygen species, two hallmarks of cancer. We further demonstrate that combination of an SLC8A1 inhibitor and a mitochondria-targeted antioxidant promotes enhanced cell death efficacy in LKB1- and SIK2-negative uveal melanoma cells compared to control cells. Our study also identified an LKB1-loss gene signature for the survival prognostic of patients with uveal melanoma that may be also predictive of response to the therapy combination. Our data thus identify not only metabolic vulnerabilities but also new prognostic markers, thereby providing a therapeutic strategy for particular subtypes of metastatic uveal melanoma.


Assuntos
Melanoma , Neoplasias Uveais , Humanos , Cálcio , Proliferação de Células , Melanoma/tratamento farmacológico , Espécies Reativas de Oxigênio , Neoplasias Uveais/genética , Neoplasias Uveais/patologia
4.
Cancer Discov ; 13(12): 2632-2651, 2023 12 12.
Artigo em Inglês | MEDLINE | ID: mdl-37676642

RESUMO

TP53 mutations are frequent in esophageal squamous cell carcinoma (ESCC) and other SCCs and are associated with a proclivity for metastasis. Here, we report that colony-stimulating factor-1 (CSF-1) expression is upregulated significantly in a p53-R172H-dependent manner in metastatic lung lesions of ESCC. The p53-R172H-dependent CSF-1 signaling, through its cognate receptor CSF-1R, increases tumor cell invasion and lung metastasis, which in turn is mediated in part through Stat3 phosphorylation and epithelial-to-mesenchymal transition (EMT). In Trp53R172H tumor cells, p53 occupies the Csf-1 promoter. The Csf-1 locus is enriched with histone 3 lysine 27 acetylation (H3K27ac), which is likely permissive for fostering an interaction between bromodomain-containing domain 4 (BRD4) and p53-R172H to regulate Csf-1 transcription. Inhibition of BRD4 not only reduces tumor invasion and lung metastasis but also reduces circulating CSF-1 levels. Overall, our results establish a novel p53-R172H-dependent BRD4-CSF-1 axis that promotes ESCC lung metastasis and suggest avenues for therapeutic strategies for this difficult-to-treat disease. SIGNIFICANCE: The invasion-metastasis cascade is a recalcitrant barrier to effective cancer therapy. We establish that the p53-R172H-dependent BRD4-CSF-1 axis is a mediator of prometastatic properties, correlates with patient survival and tumor stages, and its inhibition significantly reduces tumor cell invasion and lung metastasis. This axis can be exploited for therapeutic advantage. This article is featured in Selected Articles from This Issue, p. 2489.


Assuntos
Neoplasias Esofágicas , Carcinoma de Células Escamosas do Esôfago , Neoplasias Pulmonares , Humanos , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Transição Epitelial-Mesenquimal/genética , Neoplasias Esofágicas/genética , Carcinoma de Células Escamosas do Esôfago/genética , Mutação com Ganho de Função , Regulação Neoplásica da Expressão Gênica , Neoplasias Pulmonares/genética , Fator Estimulador de Colônias de Macrófagos/genética , Fator Estimulador de Colônias de Macrófagos/metabolismo , Mutação , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo
5.
Nat Cell Biol ; 25(9): 1332-1345, 2023 09.
Artigo em Inglês | MEDLINE | ID: mdl-37605008

RESUMO

MacroH2A has established tumour suppressive functions in melanoma and other cancers, but an unappreciated role in the tumour microenvironment. Using an autochthonous, immunocompetent mouse model of melanoma, we demonstrate that mice devoid of macroH2A variants exhibit increased tumour burden compared with wild-type counterparts. MacroH2A-deficient tumours accumulate immunosuppressive monocytes and are depleted of functional cytotoxic T cells, characteristics consistent with a compromised anti-tumour response. Single cell and spatial transcriptomics identify increased dedifferentiation along the neural crest lineage of the tumour compartment and increased frequency and activation of cancer-associated fibroblasts following macroH2A loss. Mechanistically, macroH2A-deficient cancer-associated fibroblasts display increased myeloid chemoattractant activity as a consequence of hyperinducible expression of inflammatory genes, which is enforced by increased chromatin looping of their promoters to enhancers that gain H3K27ac. In summary, we reveal a tumour suppressive role for macroH2A variants through the regulation of chromatin architecture in the tumour stroma with potential implications for human melanoma.


Assuntos
Fibroblastos Associados a Câncer , Histonas , Melanoma , Animais , Camundongos , Cromatina/genética , Expressão Gênica , Histonas/genética , Melanoma/genética , Microambiente Tumoral/genética
6.
Cell Rep ; 42(6): 112560, 2023 06 27.
Artigo em Inglês | MEDLINE | ID: mdl-37267946

RESUMO

Disseminated cancer cells (DCCs) in secondary organs can remain dormant for years to decades before reactivating into overt metastasis. Microenvironmental signals leading to cancer cell chromatin remodeling and transcriptional reprogramming appear to control onset and escape from dormancy. Here, we reveal that the therapeutic combination of the DNA methylation inhibitor 5-azacytidine (AZA) and the retinoic acid receptor ligands all-trans retinoic acid (atRA) or AM80, an RARα-specific agonist, promotes stable dormancy in cancer cells. Treatment of head and neck squamous cell carcinoma (HNSCC) or breast cancer cells with AZA+atRA induces a SMAD2/3/4-dependent transcriptional program that restores transforming growth factor ß (TGF-ß)-signaling and anti-proliferative function. Significantly, either combination, AZA+atRA or AZA+AM80, strongly suppresses HNSCC lung metastasis formation by inducing and maintaining solitary DCCs in a SMAD4+/NR2F1+ non-proliferative state. Notably, SMAD4 knockdown is sufficient to drive resistance to AZA+atRA-induced dormancy. We conclude that therapeutic doses of AZA and RAR agonists may induce and/or maintain dormancy and significantly limit metastasis development.


Assuntos
Neoplasias da Mama , Transdução de Sinais , Proteína Smad4 , Carcinoma de Células Escamosas de Cabeça e Pescoço , Tretinoína , Humanos , Azacitidina/farmacologia , Neoplasias de Cabeça e Pescoço/tratamento farmacológico , Carcinoma de Células Escamosas de Cabeça e Pescoço/tratamento farmacológico , Fator de Crescimento Transformador beta/metabolismo , Tretinoína/farmacologia , Neoplasias da Mama/tratamento farmacológico , Linhagem Celular Tumoral/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos
8.
Blood Cancer Discov ; 4(4): 318-335, 2023 07 05.
Artigo em Inglês | MEDLINE | ID: mdl-37067914

RESUMO

The reprogramming of human acute myeloid leukemia (AML) cells into induced pluripotent stem cell (iPSC) lines could provide new faithful genetic models of AML, but is currently hindered by low success rates and uncertainty about whether iPSC-derived cells resemble their primary counterparts. Here we developed a reprogramming method tailored to cancer cells, with which we generated iPSCs from 15 patients representing all major genetic groups of AML. These AML-iPSCs retain genetic fidelity and produce transplantable hematopoietic cells with hallmark phenotypic leukemic features. Critically, single-cell transcriptomics reveal that, upon xenotransplantation, iPSC-derived leukemias faithfully mimic the primary patient-matched xenografts. Transplantation of iPSC-derived leukemias capturing a clone and subclone from the same patient allowed us to isolate the contribution of a FLT3-ITD mutation to the AML phenotype. The results and resources reported here can transform basic and preclinical cancer research of AML and other human cancers. SIGNIFICANCE: We report the generation of patient-derived iPSC models of all major genetic groups of human AML. These exhibit phenotypic hallmarks of AML in vitro and in vivo, inform the clonal hierarchy and clonal dynamics of human AML, and exhibit striking similarity to patient-matched primary leukemias upon xenotransplantation. See related commentary by Doulatov, p. 252. This article is highlighted in the In This Issue feature, p. 247.


Assuntos
Células-Tronco Pluripotentes Induzidas , Leucemia Mieloide Aguda , Humanos , Células-Tronco Pluripotentes Induzidas/metabolismo , Leucemia Mieloide Aguda/genética , Fenótipo , Perfilação da Expressão Gênica , Variação Genética/genética
9.
Commun Biol ; 6(1): 215, 2023 02 23.
Artigo em Inglês | MEDLINE | ID: mdl-36823213

RESUMO

Considerable efforts have been made to characterize active enhancer elements, which can be annotated by accessible chromatin and H3 lysine 27 acetylation (H3K27ac). However, apart from poised enhancers that are observed in early stages of development and putative silencers, the functional significance of cis-regulatory elements lacking H3K27ac is poorly understood. Here we show that macroH2A histone variants mark a subset of enhancers in normal and cancer cells, which we coined 'macro-Bound Enhancers', that modulate enhancer activity. We find macroH2A variants localized at enhancer elements that are devoid of H3K27ac in a cell type-specific manner, indicating a role for macroH2A at inactive enhancers to maintain cell identity. In following, reactivation of macro-bound enhancers is associated with oncogenic programs in breast cancer and their repressive role is correlated with the activity of macroH2A2 as a negative regulator of BRD4 chromatin occupancy. Finally, through single cell epigenomic profiling of normal mammary stem cells derived from mice, we show that macroH2A deficiency facilitates increased activity of transcription factors associated with stem cell activity.


Assuntos
Proteínas Nucleares , Fatores de Transcrição , Camundongos , Animais , Proteínas Nucleares/metabolismo , Fatores de Transcrição/genética , Reprogramação Celular/genética , Elementos Facilitadores Genéticos , Cromatina/genética
10.
Sci Adv ; 8(48): eabo0876, 2022 12 02.
Artigo em Inglês | MEDLINE | ID: mdl-36459552

RESUMO

MacroH2A variants have been linked to inhibition of metastasis through incompletely understood mechanisms. Here, we reveal that solitary dormant disseminated cancer cells (DCCs) display increased levels of macroH2A variants in head and neck squamous cell carcinoma PDX in vivo models and patient samples compared to proliferating primary or metastatic lesions. We demonstrate that dormancy-inducing transforming growth factor-ß2 and p38α/ß pathways up-regulate macroH2A expression and that macroH2A variant overexpression is sufficient to induce DCC dormancy and suppress metastasis in vivo. Notably, inducible expression of the macroH2A2 variant in vivo suppresses metastasis via a reversible growth arrest of DCCs. This state does not require the dormancy-regulating transcription factors DEC2 and NR2F1; instead, transcriptomic analysis reveals that macroH2A2 overexpression inhibits cell cycle and oncogenic signaling programs, while up-regulating dormancy and senescence-associated inflammatory cytokines. We conclude that the macroH2A2-enforced dormant phenotype results from tapping into transcriptional programs of both quiescence and senescence to limit metastatic outgrowth.


Assuntos
Neoplasias de Cabeça e Pescoço , Histonas , Humanos , Carcinogênese , Divisão Celular , Ciclo Celular , Neoplasias de Cabeça e Pescoço/genética
11.
Nat Commun ; 13(1): 6041, 2022 10 17.
Artigo em Inglês | MEDLINE | ID: mdl-36253360

RESUMO

Tumors exhibit enhancer reprogramming compared to normal tissue. The etiology is largely attributed to cell-intrinsic genomic alterations. Here, using freshly resected primary CRC tumors and patient-matched adjacent normal colon, we find divergent epigenetic landscapes between CRC tumors and cell lines. Intriguingly, this phenomenon extends to highly recurrent aberrant super-enhancers gained in CRC over normal. We find one such super-enhancer activated in epithelial cancer cells due to surrounding inflammation in the tumor microenvironment. We restore this super-enhancer and its expressed gene, PDZK1IP1, following treatment with cytokines or xenotransplantation into nude mice, thus demonstrating cell-extrinsic etiology. We demonstrate mechanistically that PDZK1IP1 enhances the reductive capacity CRC cancer cells via the pentose phosphate pathway. We show this activation enables efficient growth under oxidative conditions, challenging the previous notion that PDZK1IP1 acts as a tumor suppressor in CRC. Collectively, these observations highlight the significance of epigenomic profiling on primary specimens.


Assuntos
Neoplasias Colorretais , Microambiente Tumoral , Animais , Carcinogênese/genética , Linhagem Celular Tumoral , Proliferação de Células/genética , Neoplasias Colorretais/patologia , Citocinas/metabolismo , Elementos Facilitadores Genéticos/genética , Regulação Neoplásica da Expressão Gênica , Camundongos , Camundongos Nus , Microambiente Tumoral/genética
12.
Cell Rep ; 39(1): 110637, 2022 04 05.
Artigo em Inglês | MEDLINE | ID: mdl-35385731

RESUMO

ARID2 is the most recurrently mutated SWI/SNF complex member in melanoma; however, its tumor-suppressive mechanisms in the context of the chromatin landscape remain to be elucidated. Here, we model ARID2 deficiency in melanoma cells, which results in defective PBAF complex assembly with a concomitant genomic redistribution of the BAF complex. Upon ARID2 depletion, a subset of PBAF and shared BAF-PBAF-occupied regions displays diminished chromatin accessibility and associated gene expression, while BAF-occupied enhancers gain chromatin accessibility and expression of genes linked to the process of invasion. As a function of altered accessibility, the genomic occupancy of melanoma-relevant transcription factors is affected and significantly correlates with the observed transcriptional changes. We further demonstrate that ARID2-deficient cells acquire the ability to colonize distal organs in multiple animal models. Taken together, our results reveal a role for ARID2 in mediating BAF and PBAF subcomplex chromatin dynamics with consequences for melanoma metastasis.


Assuntos
Proteínas Cromossômicas não Histona , Melanoma , Fatores de Transcrição , Animais , Cromatina , Montagem e Desmontagem da Cromatina , Regulação da Expressão Gênica , Humanos , Melanoma/genética , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo
13.
Nat Commun ; 9(1): 3440, 2018 08 24.
Artigo em Inglês | MEDLINE | ID: mdl-30143629

RESUMO

While multiple mechanisms of BRAFV600-mutant melanoma resistance to targeted MAPK signaling inhibitors (MAPKi) have been reported, the epigenetic regulation of this process remains undetermined. Here, using a CRISPR-Cas9 screen targeting chromatin regulators, we discover that haploinsufficiency of the histone deacetylase SIRT6 allows melanoma cell persistence in the presence of MAPKi. Haploinsufficiency, but not complete loss of SIRT6 promotes IGFBP2 expression via increased chromatin accessibility, H3K56 acetylation at the IGFBP2 locus, and consequent activation of the IGF-1 receptor (IGF-1R) and downstream AKT signaling. Combining a clinically applicable IGF-1Ri with BRAFi overcomes resistance of SIRT6 haploinsufficient melanoma cells in vitro and in vivo. Using matched melanoma samples derived from patients receiving dabrafenib + trametinib, we identify IGFBP2 as a potential biomarker for MAPKi resistance. Our study has not only identified an epigenetic mechanism of drug resistance, but also provides insights into a combinatorial therapy that may overcome resistance to standard-of-care therapy for BRAFV600-mutant melanoma patients.


Assuntos
Haploinsuficiência/fisiologia , Melanoma/tratamento farmacológico , Melanoma/metabolismo , Inibidores de Proteínas Quinases/uso terapêutico , Proteínas Proto-Oncogênicas B-raf/metabolismo , Receptor IGF Tipo 1/metabolismo , Sirtuínas/metabolismo , Animais , Apoptose/efeitos dos fármacos , Apoptose/genética , Linhagem Celular , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/genética , Imunoprecipitação da Cromatina , Dano ao DNA/efeitos dos fármacos , Dano ao DNA/genética , Feminino , Haploinsuficiência/genética , Humanos , Melanoma/genética , Camundongos Nus , Proteínas Proto-Oncogênicas B-raf/genética , Receptor IGF Tipo 1/genética , Transdução de Sinais/genética , Transdução de Sinais/fisiologia , Sirtuínas/genética
14.
Proc Natl Acad Sci U S A ; 108(52): 21069-74, 2011 Dec 27.
Artigo em Inglês | MEDLINE | ID: mdl-22106309

RESUMO

During cell division, the activation of glycolysis is tightly regulated by the action of two ubiquitin ligases, anaphase-promoting complex/cyclosome-Cdh1 (APC/C-Cdh1) and SKP1/CUL-1/F-box protein-ß-transducin repeat-containing protein (SCF-ß-TrCP), which control the transient appearance and metabolic activity of the glycolysis-promoting enzyme 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase, isoform 3 (PFKFB3). We now demonstrate that the breakdown of PFKFB3 during S phase occurs specifically via a distinct residue (S(273)) within the conserved recognition site for SCF-ß-TrCP. Glutaminase 1 (GLS1), the first enzyme in glutaminolysis, is also targeted for destruction by APC/C-Cdh1 and, like PFKFB3, accumulates after the activity of this ubiquitin ligase decreases in mid-to-late G1. However, our results show that GLS1 differs from PFKFB3 in that its recognition by APC/C-Cdh1 requires the presence of both a Lys-Glu-Asn box (KEN box) and a destruction box (D box) rather than a KEN box alone. Furthermore, GLS1 is not a substrate for SCF-ß-TrCP and is not degraded until cells progress from S to G2/M. The presence of PFKFB3 and GLS1 coincides with increases in generation of lactate and in utilization of glutamine, respectively. The contrasting posttranslational regulation of PFKFB3 and GLS1, which we have verified by studies of ubiquitination and protein stability, suggests the different roles of glucose and glutamine at distinct stages in the cell cycle. Indeed, experiments in which synchronized cells were deprived of either of these substrates show that both glucose and glutamine are required for progression through the restriction point in mid-to-late G1, whereas glutamine is the only substrate essential for the progression through S phase into cell division.


Assuntos
Divisão Celular/fisiologia , Glutaminase/metabolismo , Glicólise/fisiologia , Fosfofrutoquinase-2/metabolismo , Processamento de Proteína Pós-Traducional/fisiologia , Proteínas Ligases SKP Culina F-Box/metabolismo , Complexos Ubiquitina-Proteína Ligase/metabolismo , Ciclossomo-Complexo Promotor de Anáfase , Primers do DNA/genética , Citometria de Fluxo , Glucose/metabolismo , Glutamina/metabolismo , Células HeLa , Humanos , Immunoblotting , Plasmídeos/genética , Ubiquitinação
15.
Proc Natl Acad Sci U S A ; 108(13): 5278-83, 2011 Mar 29.
Artigo em Inglês | MEDLINE | ID: mdl-21402913

RESUMO

During cell proliferation, the abundance of the glycolysis-promoting enzyme, 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase, isoform 3 (PFKFB3), is controlled by the ubiquitin ligase APC/C-Cdh1 via a KEN box. We now demonstrate in synchronized HeLa cells that PFKFB3, which appears in mid-to-late G1, is essential for cell division because its silencing prevents progression into S phase. In cells arrested by glucose deprivation, progression into S phase after replacement of glucose occurs only when PFKFB3 is present or is substituted by the downstream glycolytic enzyme 6-phosphofructo-1-kinase. PFKFB3 ceases to be detectable during late G1/S despite the absence of Cdh1; this disappearance is prevented by proteasomal inhibition. PFKFB3 contains a DSG box and is therefore a potential substrate for SCF-ß-TrCP, a ubiquitin ligase active during S phase. In synchronized HeLa cells transfected with PFKFB3 mutated in the KEN box, the DSG box, or both, we established the breakdown routes of the enzyme at different stages of the cell cycle and the point at which glycolysis is enhanced. Thus, the presence of PFKFB3 is tightly controlled to ensure the up-regulation of glycolysis at a specific point in G1. We suggest that this up-regulation of glycolysis and its associated events represent the nutrient-sensitive restriction point in mammalian cells.


Assuntos
Ciclo Celular/fisiologia , Glicólise/fisiologia , Fosfofrutoquinase-2/metabolismo , Proteínas Ligases SKP Culina F-Box/metabolismo , Complexos Ubiquitina-Proteína Ligase/metabolismo , Sequência de Aminoácidos , Ciclossomo-Complexo Promotor de Anáfase , Proliferação de Células , Estabilidade Enzimática , Glucose/metabolismo , Células HeLa , Humanos , Ácido Láctico/metabolismo , Dados de Sequência Molecular , Fosfofrutoquinase-2/genética , Interferência de RNA
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