RESUMO
The AP-1 transcription factor c-Jun is required for Ras-driven tumorigenesis in many tissues and is considered as a classical proto-oncogene. To determine the requirement for c-Jun in a mouse model of K-RasG12D-induced lung adenocarcinoma, we inducibly deleted c-Jun in the adult lung. Surprisingly, we found that inactivation of c-Jun, or mutation of its JNK phosphorylation sites, actually increased lung tumor burden. Mechanistically, we found that protein levels of the Jun family member JunD were increased in the absence of c-Jun. In c-Jun-deficient cells, JunD phosphorylation was increased, and expression of a dominant-active JNKK2-JNK1 transgene further increased lung tumor formation. Strikingly, deletion of JunD completely abolished Ras-driven lung tumorigenesis. This work identifies JunD, not c-Jun, as the crucial substrate of JNK signaling and oncogene required for Ras-induced lung cancer.
Assuntos
Adenocarcinoma de Pulmão , Carcinogênese , Neoplasias Pulmonares , Proteínas Proto-Oncogênicas c-jun/metabolismo , Proteínas ras/metabolismo , Adenocarcinoma de Pulmão/genética , Adenocarcinoma de Pulmão/metabolismo , Animais , Carcinogênese/genética , Carcinogênese/metabolismo , Regulação Neoplásica da Expressão Gênica/genética , Inativação Gênica , Genes jun/genética , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , MAP Quinase Quinase 7/genética , MAP Quinase Quinase 7/metabolismo , Sistema de Sinalização das MAP Quinases , Camundongos , Fosforilação , Proteínas Proto-Oncogênicas c-jun/genética , Fator de Transcrição AP-1/metabolismoRESUMO
RAC1 P29 is the third most commonly mutated codon in human cutaneous melanoma, after BRAF V600 and NRAS Q61. Here, we study the role of RAC1P29S in melanoma development and reveal that RAC1P29S activates PAK, AKT, and a gene expression program initiated by the SRF/MRTF transcriptional pathway, which results in a melanocytic to mesenchymal phenotypic switch. Mice with ubiquitous expression of RAC1P29S from the endogenous locus develop lymphoma. When expressed only in melanocytes, RAC1P29S cooperates with oncogenic BRAF or with NF1-loss to promote tumorigenesis. RAC1P29S also drives resistance to BRAF inhibitors, which is reversed by SRF/MRTF inhibitors. These findings establish RAC1P29S as a promoter of melanoma initiation and mediator of therapy resistance, while identifying SRF/MRTF as a potential therapeutic target.
Assuntos
Transformação Celular Neoplásica/genética , Resistencia a Medicamentos Antineoplásicos/genética , Transição Epitelial-Mesenquimal/genética , Melanoma/etiologia , Melanoma/patologia , Mutação , Proteínas rac1 de Ligação ao GTP/genética , Alelos , Substituição de Aminoácidos , Animais , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/genética , Modelos Animais de Doenças , Feminino , Expressão Gênica , Humanos , Masculino , Melanócitos/metabolismo , Melanoma/mortalidade , Melanoma/terapia , Camundongos , Camundongos Transgênicos , Modelos Biológicos , Prognóstico , Inibidores de Proteínas Quinases/farmacologia , Inibidores de Proteínas Quinases/uso terapêutico , Proteínas Proto-Oncogênicas B-raf/genética , Fator de Resposta Sérica , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Colorectal cancer (CRC) patients develop recurrence after chemotherapy owing to the survival of stem cell-like cells referred to as cancer stem-like cells (CSCs). The origin of CSCs is linked to the epithelial-mesenchymal transition (EMT) process. Currently, it remains poorly understood how EMT programmes enable CSCs residing in the tumour microenvironment to escape the effects of chemotherapy. This study identifies a key molecular pathway that is responsible for the formation of drug-resistant CSC populations. Using a modified yeast-2-hybrid system and 2D gel-based proteomics methods, we show that the E3-ubiquitin ligase FBXW7 directly binds and degrades the EMT-inducing transcription factor ZEB2 in a phosphorylation-dependent manner. Loss of FBXW7 induces an EMT that can be effectively reversed by knockdown of ZEB2. The FBXW7-ZEB2 axis regulates such important cancer cell features, as stemness/dedifferentiation, chemoresistance and cell migration in vitro, ex vivo and in animal models of metastasis. High expression of ZEB2 in cancer tissues defines the reduced ZEB2 expression in the cancer-associated stroma in patients and in murine intestinal organoids, demonstrating a tumour-stromal crosstalk that modulates a niche and EMT activation. Our study thus uncovers a new molecular mechanism, by which the CRC cells display differences in resistance to chemotherapy and metastatic potential.
RESUMO
RAS family GTPases contribute directly to the regulation of type I phosphoinositide 3-kinases (PI3Ks) via RAS-binding domains in the PI3K catalytic p110 subunits. Disruption of this domain of p110α impairs RAS-mutant-oncogene-driven tumor formation and maintenance. Here, we test the effect of blocking the interaction of RAS with p110α on epidermal growth factor receptor (EGFR)-mutant-driven lung tumorigenesis. Disrupting the RAS-PI3K interaction inhibits activation of both AKT and RAC1 in EGFR-mutant lung cancer cells, leading to reduced growth and survival, and inhibits EGFR-mutant-induced tumor onset and promotes major regression of established tumors in an autochthonous mouse model of EGFR-mutant-induced lung adenocarcinoma. The RAS-PI3K interaction is thus an important signaling node and potential therapeutic target in EGFR-mutant lung cancer, even though RAS oncogenes are not themselves mutated in this setting, suggesting different strategies for tackling tyrosine kinase inhibitor resistance in lung cancer.
Assuntos
Receptores ErbB/genética , Neoplasias Pulmonares/genética , Mutação/genética , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas ras/metabolismo , Animais , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Modelos Animais de Doenças , Fator de Crescimento Epidérmico/farmacologia , Receptores ErbB/metabolismo , Humanos , Neoplasias Pulmonares/patologia , Camundongos Endogâmicos C57BL , Fosfatidilinositol 3-Quinases/química , Ligação Proteica/efeitos dos fármacos , Domínios ProteicosRESUMO
Squamous cell carcinoma (SCC) can progress to malignant metastatic cancer, including an aggressive subtype known as spindle cell carcinoma (spSCC). spSCC formation involves epithelial-to-mesenchymal transition (EMT), yet the molecular basis of this event remains unknown. The transcriptional co-activator YAP undergoes recurrent amplification in human SCC and overexpression of YAP drives SCC formation in mice. Here, we show that human spSCC tumours also feature strong nuclear localisation of YAP and overexpression of activated YAP (NLS-YAP-5SA) with Keratin-5 (K5-CreERt) is sufficient to induce rapid formation of both SCC and spSCC in mice. spSCC tumours arise at sites of epithelial scratch wounding, where tumour-initiating epithelial cells undergo EMT to generate spSCC. Expression of the EMT transcription factor ZEB1 arises upon wounding and is a defining characteristic of spSCC in mice and humans. Thus, the wound healing response synergises with YAP to drive metaplastic transformation of SCC to spSCC.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Carcinogênese/metabolismo , Carcinogênese/patologia , Carcinoma de Células Escamosas/metabolismo , Carcinoma de Células Escamosas/patologia , Progressão da Doença , Fosfoproteínas/metabolismo , Neoplasias Cutâneas/metabolismo , Neoplasias Cutâneas/patologia , Animais , Núcleo Celular/metabolismo , Epiderme/patologia , Transição Epitelial-Mesenquimal , Humanos , Camundongos , Fatores de Transcrição , Proteínas de Sinalização YAP , Homeobox 1 de Ligação a E-box em Dedo de Zinco/metabolismoRESUMO
Wnt/ß-catenin signaling plays a critical role during development of both normal and malignant colorectal cancer tissues. Phosphorylation of ß-catenin protein alters its trafficking and function. Such conventional allosteric regulation usually involves a highly specialized set of molecular interactions, which may specifically turn on a particular cell phenotype. This study identifies a novel transcription modulator with an FLYWCH/Zn-finger DNA-binding domain, called "FLYWCH1." Using a modified yeast-2-hybrid based Ras-Recruitment system, it is demonstrated that FLYWCH1 directly binds to unphosphorylated (nuclear) ß-catenin efficiently suppressing the transcriptional activity of Wnt/ß-catenin signaling that cannot be rescued by TCF4. FLYWCH1 rearranges the transcriptional activity of ß-catenin/TCF4 to selectively block the expression of specific downstream genes associated with colorectal cancer cell migration and morphology, including ZEB1, EPHA4, and E-cadherin. Accordingly, overexpression of FLYWCH1 reduces cell motility and increases cell attachment. The expression of FLYWCH1 negatively correlates with the expression level of ZEB1 and EPHA4 in normal versus primary and metastatic colorectal cancer tissues in patients. Thus, FLYWCH1 antagonizes ß-catenin/TCF4 signaling during cell polarity/migration in colorectal cancer. IMPLICATIONS: This study uncovers a new molecular mechanism by which FLYWCH1 with a possible tumor suppressive role represses ß-catenin-induced ZEB1 and increases cadherin-mediated cell attachment preventing colorectal cancer metastasis.
Assuntos
Neoplasias Colorretais/metabolismo , Proteínas de Ligação a DNA/metabolismo , Via de Sinalização Wnt , beta Catenina/metabolismo , Antígenos CD/metabolismo , Caderinas/metabolismo , Linhagem Celular Tumoral , Movimento Celular , Proteínas de Ligação a DNA/química , Regulação para Baixo , Regulação Neoplásica da Expressão Gênica , Células HCT116 , Humanos , Análise Serial de Tecidos , Homeobox 1 de Ligação a E-box em Dedo de Zinco/metabolismo , Dedos de ZincoRESUMO
The tumor suppressor FBW7 targets oncoproteins such as c-MYC for ubiquitylation and is mutated in several human cancers. We noted that in a substantial percentage of colon cancers, FBW7 protein is undetectable despite the presence of FBW7 mRNA. To understand the molecular mechanism of FBW7 regulation in these cancers, we employed proteomics and identified the deubiquitinase (DUB) USP9X as an FBW7 interactor. USP9X antagonized FBW7 ubiquitylation, and Usp9x deletion caused Fbw7 destabilization. Mice lacking Usp9x in the gut showed reduced secretory cell differentiation and increased progenitor proliferation, phenocopying Fbw7 loss. In addition, Usp9x inactivation impaired intestinal regeneration and increased tumor burden in colitis-associated intestinal cancer. c-Myc heterozygosity abrogated increased progenitor proliferation and tumor burden in Usp9x-deficient mice, suggesting that Usp9x suppresses tumor formation by regulating Fbw7 protein stability and thereby reducing c-Myc. Thus, we identify a tumor suppressor mechanism in the mammalian intestine that arises from the posttranslational regulation of FBW7 by USP9X independent of somatic FBW7 mutations.
Assuntos
Neoplasias Colorretais/enzimologia , Endopeptidases/biossíntese , Proteína 7 com Repetições F-Box-WD/biossíntese , Regulação Enzimológica da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Proteínas Supressoras de Tumor/biossíntese , Ubiquitina Tiolesterase/biossíntese , Animais , Neoplasias Colorretais/genética , Neoplasias Colorretais/patologia , Endopeptidases/genética , Proteína 7 com Repetições F-Box-WD/genética , Células HCT116 , Humanos , Camundongos , Camundongos Mutantes , Muramidase , Mutação , Fragmentos de Peptídeos , Proteínas Proto-Oncogênicas c-myc/genética , Proteínas Proto-Oncogênicas c-myc/metabolismo , Proteínas Supressoras de Tumor/genética , Ubiquitina Tiolesterase/genéticaRESUMO
The immunosuppressive protein PD-L1 is upregulated in many cancers and contributes to evasion of the host immune system. The relative importance of the tumor microenvironment and cancer cell-intrinsic signaling in the regulation of PD-L1 expression remains unclear. We report that oncogenic RAS signaling can upregulate tumor cell PD-L1 expression through a mechanism involving increases in PD-L1 mRNA stability via modulation of the AU-rich element-binding protein tristetraprolin (TTP). TTP negatively regulates PD-L1 expression through AU-rich elements in the 3' UTR of PD-L1 mRNA. MEK signaling downstream of RAS leads to phosphorylation and inhibition of TTP by the kinase MK2. In human lung and colorectal tumors, RAS pathway activation is associated with elevated PD-L1 expression. In vivo, restoration of TTP expression enhances anti-tumor immunity dependent on degradation of PD-L1 mRNA. We demonstrate that RAS can drive cell-intrinsic PD-L1 expression, thus presenting therapeutic opportunities to reverse the innately immunoresistant phenotype of RAS mutant cancers.
Assuntos
Antígeno B7-H1/imunologia , Neoplasias Colorretais/imunologia , Regulação Neoplásica da Expressão Gênica , Neoplasias Pulmonares/imunologia , Proteínas Proto-Oncogênicas p21(ras)/imunologia , Tristetraprolina/imunologia , Evasão Tumoral , Animais , Antígeno B7-H1/genética , Linhagem Celular Tumoral , Neoplasias Colorretais/genética , Neoplasias Colorretais/patologia , Células Epiteliais/imunologia , Células Epiteliais/patologia , Feminino , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/genética , Peptídeos e Proteínas de Sinalização Intracelular/imunologia , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , MAP Quinase Quinase Quinases/genética , MAP Quinase Quinase Quinases/imunologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Transplante de Neoplasias , Ligação Proteica , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/imunologia , Proteínas Proto-Oncogênicas p21(ras)/genética , Clivagem do RNA , Estabilidade de RNA , RNA Mensageiro/genética , RNA Mensageiro/imunologia , Transdução de Sinais , Tristetraprolina/genéticaRESUMO
The cell of origin of pancreatic ductal adenocarcinoma (PDAC) has been controversial. Here, we show that identical oncogenic drivers trigger PDAC originating from both ductal and acinar cells with similar histology but with distinct pathophysiology and marker expression dependent on cell of origin. Whereas acinar-derived tumors exhibited low AGR2 expression and were preceded by pancreatic intraepithelial neoplasias (PanINs), duct-derived tumors displayed high AGR2 and developed independently of a PanIN stage via non-mucinous lesions. Using orthotopic transplantation and chimera experiments, we demonstrate that PanIN-like lesions can be induced by PDAC as bystanders in adjacent healthy tissues, explaining the co-existence of mucinous and non-mucinous lesions and highlighting the need to distinguish between true precursor PanINs and PanIN-like bystander lesions. Our results suggest AGR2 as a tool to stratify PDAC according to cell of origin, highlight that not all PanIN-like lesions are precursors of PDAC, and add an alternative progression route to the current model of PDAC development.
Assuntos
Células Acinares/patologia , Carcinogênese , Carcinoma Ductal Pancreático/patologia , Ductos Pancreáticos/patologia , Neoplasias Pancreáticas/patologia , Células Acinares/metabolismo , Animais , Carcinoma Ductal Pancreático/metabolismo , Células Cultivadas , Feminino , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Mucoproteínas/genética , Mucoproteínas/metabolismo , Proteínas Oncogênicas , Ductos Pancreáticos/metabolismo , Neoplasias Pancreáticas/metabolismoRESUMO
The myocardin-related transcription factor/serum response factor (MRTF/SRF) pathway represents a promising therapeutic target to prevent fibrosis. We have tested the effects of new pharmacological inhibitors of MRTF/SRF signalling in a preclinical model of fibrosis. CCG-222740, a novel MRTF/SRF inhibitor, markedly decreased SRF reporter gene activity and showed a greater inhibitory effect on MRTF/SRF target genes than the previously described MRTF-A inhibitor CCG-203971. CCG-222740 was also five times more potent, with an IC50 of 5 µM, in a fibroblast-mediated collagen contraction assay, was less cytotoxic, and a more potent inhibitor of alpha-smooth muscle actin protein expression than CCG-203971. Local delivery of CCG-222740 and CCG-203971 in a validated and clinically relevant rabbit model of scar tissue formation after glaucoma filtration surgery increased the long-term success of the surgery by 67% (P < 0.0005) and 33% (P < 0.01), respectively, and significantly decreased fibrosis and scarring histologically. Unlike mitomycin-C, neither CCG-222740 nor CCG-203971 caused any detectable epithelial toxicity or systemic side effects with very low drug levels measured in the aqueous, vitreous, and serum. We conclude that inhibitors of MRTF/SRF-regulated gene transcription such as CCG-222740, potentially represent a new therapeutic strategy to prevent scar tissue formation in the eye and other tissues.
Assuntos
Cicatriz/metabolismo , Cicatriz/patologia , Fator de Resposta Sérica/antagonistas & inibidores , Fator de Resposta Sérica/metabolismo , Transativadores/antagonistas & inibidores , Transativadores/metabolismo , Animais , Células Cultivadas , Cicatriz/prevenção & controle , Colágeno/metabolismo , Modelos Animais de Doenças , Avaliação Pré-Clínica de Medicamentos , Matriz Extracelular , Feminino , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Fibrose , Humanos , Coelhos , Transdução de Sinais/efeitos dos fármacosRESUMO
Chromosomal instability (CIN) contributes to cancer evolution, intratumor heterogeneity, and drug resistance. CIN is driven by chromosome segregation errors and a tolerance phenotype that permits the propagation of aneuploid genomes. Through genomic analysis of colorectal cancers and cell lines, we find frequent loss of heterozygosity and mutations in BCL9L in aneuploid tumors. BCL9L deficiency promoted tolerance of chromosome missegregation events, propagation of aneuploidy, and genetic heterogeneity in xenograft models likely through modulation of Wnt signaling. We find that BCL9L dysfunction contributes to aneuploidy tolerance in both TP53-WT and mutant cells by reducing basal caspase-2 levels and preventing cleavage of MDM2 and BID. Efforts to exploit aneuploidy tolerance mechanisms and the BCL9L/caspase-2/BID axis may limit cancer diversity and evolution.
Assuntos
Aneuploidia , Caspase 2/fisiologia , Neoplasias Colorretais/genética , Cisteína Endopeptidases/fisiologia , Proteínas de Ligação a DNA/fisiologia , Fatores de Transcrição/fisiologia , Idoso , Idoso de 80 Anos ou mais , Animais , Proteína Agonista de Morte Celular de Domínio Interatuante com BH3/fisiologia , Caspase 2/análise , Segregação de Cromossomos , Cisteína Endopeptidases/análise , Proteínas de Ligação a DNA/genética , Células HCT116 , Humanos , Camundongos , Pessoa de Meia-Idade , Mutação , Proteínas Proto-Oncogênicas c-mdm2/fisiologia , Fatores de Transcrição/genética , Proteína Supressora de Tumor p53/fisiologiaRESUMO
Microvascular endothelial cells (ECs) display a high degree of phenotypic and functional heterogeneity among different organs. Organ-specific ECs control their tissue microenvironment by angiocrine factors in health and disease. Liver sinusoidal endothelial cells (LSECs) are uniquely differentiated to fulfill important organ-specific functions in development, under homeostatic conditions, and in regeneration and liver pathology. Recently, Bmp2 has been identified by us as an organ-specific angiokine derived from LSECs. To study angiocrine Bmp2 signaling in the liver, we conditionally deleted Bmp2 in LSECs using EC subtype-specific Stab2-Cre mice. Genetic inactivation of hepatic angiocrine Bmp2 signaling in Stab2-Cre;Bmp2fl/fl (Bmp2LSECKO) mice caused massive iron overload in the liver and increased serum iron levels and iron deposition in several organs similar to classic hereditary hemochromatosis. Iron overload was mediated by decreased hepatic expression of hepcidin, a key regulator of iron homeostasis. Thus, angiocrine Bmp2 signaling within the hepatic vascular niche represents a constitutive pathway indispensable for iron homeostasis in vivo that is nonredundant with Bmp6. Notably, we demonstrate that organ-specific angiocrine signaling is essential not only for the homeostasis of the respective organ but also for the homeostasis of the whole organism.
Assuntos
Proteína Morfogenética Óssea 2/genética , Células Endoteliais/metabolismo , Hemocromatose/genética , Hepcidinas/genética , Homeostase/genética , Ferro/metabolismo , Fígado/metabolismo , Animais , Proteína Morfogenética Óssea 2/deficiência , Proteína Morfogenética Óssea 6/genética , Proteína Morfogenética Óssea 6/metabolismo , Capilares/metabolismo , Capilares/patologia , Moléculas de Adesão Celular Neuronais/genética , Moléculas de Adesão Celular Neuronais/metabolismo , Células Endoteliais/patologia , Feminino , Regulação da Expressão Gênica , Hemocromatose/metabolismo , Hemocromatose/patologia , Hepatócitos/metabolismo , Hepatócitos/patologia , Hepcidinas/metabolismo , Integrases/genética , Integrases/metabolismo , Fígado/irrigação sanguínea , Fígado/patologia , Masculino , Camundongos , Camundongos Transgênicos , Comunicação Parácrina , Transdução de Sinais , Transcrição GênicaRESUMO
The mammary gland is composed of a complex cellular hierarchy with unusual postnatal plasticity. The identities of stem/progenitor cell populations, as well as tumour-initiating cells that give rise to breast cancer, are incompletely understood. Here we show that Lgr6 marks rare populations of cells in both basal and luminal mammary gland compartments in mice. Lineage tracing analysis showed that Lgr6+ cells are unipotent progenitors, which expand clonally during puberty but diminish in adulthood. In pregnancy or following stimulation with ovarian hormones, adult Lgr6+ cells regained proliferative potency and their progeny formed alveoli over repeated pregnancies. Oncogenic mutations in Lgr6+ cells resulted in expansion of luminal cells, culminating in mammary gland tumours. Conversely, depletion of Lgr6+ cells in the MMTV-PyMT model of mammary tumorigenesis significantly impaired tumour growth. Thus, Lgr6 marks mammary gland progenitor cells that can initiate tumours, and cells of luminal breast tumours required for efficient tumour maintenance.
Assuntos
Neoplasias da Mama/patologia , Glândulas Mamárias Animais/patologia , Neoplasias Mamárias Experimentais/patologia , Receptores Acoplados a Proteínas G/metabolismo , Células-Tronco/patologia , Alelos , Animais , Animais Recém-Nascidos , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/genética , Carcinogênese/patologia , Linhagem da Célula , Proliferação de Células , Células Clonais , Intervalo Livre de Doença , Resistencia a Medicamentos Antineoplásicos , Feminino , Regulação Neoplásica da Expressão Gênica , Homeostase , Hormônios/farmacologia , Humanos , Glândulas Mamárias Animais/crescimento & desenvolvimento , Neoplasias Mamárias Experimentais/genética , Camundongos , Células-Tronco Neoplásicas/metabolismo , Células-Tronco Neoplásicas/patologia , Gravidez , Células-Tronco/metabolismo , Regulação para CimaRESUMO
Tumors comprise functionally diverse subpopulations of cells with distinct proliferative potential. Here, we show that dynamic epigenetic states defined by the linker histone H1.0 determine which cells within a tumor can sustain the long-term cancer growth. Numerous cancer types exhibit high inter- and intratumor heterogeneity of H1.0, with H1.0 levels correlating with tumor differentiation status, patient survival, and, at the single-cell level, cancer stem cell markers. Silencing of H1.0 promotes maintenance of self-renewing cells by inducing derepression of megabase-sized gene domains harboring downstream effectors of oncogenic pathways. Self-renewing epigenetic states are not stable, and reexpression of H1.0 in subsets of tumor cells establishes transcriptional programs that restrict cancer cells' long-term proliferative potential and drive their differentiation. Our results uncover epigenetic determinants of tumor-maintaining cells.
Assuntos
Epigênese Genética , Regulação Neoplásica da Expressão Gênica , Heterogeneidade Genética , Histonas/genética , Neoplasias/genética , Neoplasias/patologia , Adenina/química , Linhagem Celular Tumoral , DNA/química , Metilação de DNA , Elementos Facilitadores Genéticos , Técnicas de Silenciamento de Genes , Humanos , Neoplasias/mortalidade , Nucleossomos/metabolismo , RNA Interferente Pequeno/genética , Timina/químicaRESUMO
Many body surfaces harbor organ-specific γδ T cell compartments that contribute to tissue integrity. Thus, murine dendritic epidermal T cells (DETCs) uniquely expressing T cell receptor (TCR)-Vγ5 chains protect from cutaneous carcinogens. The DETC repertoire is shaped by Skint1, a butyrophilin-like (Btnl) gene expressed specifically by thymic epithelial cells and suprabasal keratinocytes. However, the generality of this mechanism has remained opaque, since neither Skint1 nor DETCs are evolutionarily conserved. Here, Btnl1 expressed by murine enterocytes is shown to shape the local TCR-Vγ7(+) γδ compartment. Uninfluenced by microbial or food antigens, this activity evokes the developmental selection of TCRαß(+) repertoires. Indeed, Btnl1 and Btnl6 jointly induce TCR-dependent responses specifically in intestinal Vγ7(+) cells. Likewise, human gut epithelial cells express BTNL3 and BTNL8 that jointly induce selective TCR-dependent responses of human colonic Vγ4(+) cells. Hence, a conserved mechanism emerges whereby epithelia use organ-specific BTNL/Btnl genes to shape local T cell compartments.
Assuntos
Butirofilinas/imunologia , Mucosa Intestinal/imunologia , Receptores de Antígenos de Linfócitos T gama-delta/imunologia , Linfócitos T/imunologia , Animais , Butirofilinas/genética , Técnicas de Inativação de Genes , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Timo/imunologiaRESUMO
BACKGROUND & AIMS: Pancreatitis is the most important risk factor for pancreatic ductal adenocarcinoma (PDAC). Pancreatitis predisposes to PDAC because it induces a process of acinar cell reprogramming known as acinar-to-ductal metaplasia (ADM)-a precursor of pancreatic intraepithelial neoplasia lesions that can progress to PDAC. Mutations in KRAS are found at the earliest stages of pancreatic tumorigenesis, and it appears to be a gatekeeper to cancer progression. We investigated how mutations in KRAS cooperate with pancreatitis to promote pancreatic cancer progression in mice. METHODS: We generated mice carrying conditional alleles of Yap1 and Taz and disrupted Yap1 and Taz using a Cre-lox recombination strategy in adult mouse pancreatic acinar cells (Yap1fl/fl;Tazfl/fl;Ela1-CreERT2). We crossed these mice with LSL-KrasG12D mice, which express a constitutively active form of KRAS after Cre recombination. Pancreatic tumor initiation and progression were analyzed after chemically induced pancreatitis. We analyzed pancreatic tissues from patients with pancreatitis or PDAC by immunohistochemistry. RESULTS: Oncogenic activation of KRAS in normal, untransformed acinar cells in the pancreatic tissues of mice resulted in increased levels of pancreatitis-induced ADM. Expression of the constitutive active form of KRAS in this system led to activation of the transcriptional regulators YAP1 and TAZ; their function was required for pancreatitis-induced ADM in mice. The JAK-STAT3 pathway was a downstream effector of KRAS signaling via YAP1 and TAZ. YAP1 and TAZ directly mediated transcriptional activation of several genes in the JAK-STAT3 signaling pathway; this could be a mechanism by which acinar cells that express activated KRAS become susceptible to inflammation. CONCLUSIONS: We identified a mechanism by which oncogenic KRAS facilitates ADM and thereby generates the cells that initiate neoplastic progression. This process involves activation of YAP1 and TAZ in acinar cells, which up-regulate JAK-STAT3 signaling to promote development of PDAC in mice.
Assuntos
Carcinogênese/genética , Neoplasias Pancreáticas/genética , Pancreatite/complicações , Transdução de Sinais/genética , Células Acinares/metabolismo , Aciltransferases , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Animais , Proteínas de Ciclo Celular , Janus Quinases/metabolismo , Camundongos , Mutação , Pâncreas/patologia , Neoplasias Pancreáticas/etiologia , Neoplasias Pancreáticas/patologia , Pancreatite/induzido quimicamente , Pancreatite/patologia , Fosfoproteínas/metabolismo , Proteínas Proto-Oncogênicas p21(ras)/metabolismo , Fatores de Risco , Fator de Transcrição STAT3/metabolismo , Fatores de Transcrição/metabolismo , Regulação para Cima , Proteínas de Sinalização YAPRESUMO
BACKGROUND: Enhanced macromolecule biosynthesis is integral to growth and proliferation of cancer cells. Lipid biosynthesis has been predicted to be an essential process in cancer cells. However, it is unclear which enzymes within this pathway offer the best selectivity for cancer cells and could be suitable therapeutic targets. RESULTS: Using functional genomics, we identified stearoyl-CoA desaturase (SCD), an enzyme that controls synthesis of unsaturated fatty acids, as essential in breast and prostate cancer cells. SCD inhibition altered cellular lipid composition and impeded cell viability in the absence of exogenous lipids. SCD inhibition also altered cardiolipin composition, leading to the release of cytochrome C and induction of apoptosis. Furthermore, SCD was required for the generation of poly-unsaturated lipids in cancer cells grown in spheroid cultures, which resemble those found in tumour tissue. We also found that SCD mRNA and protein expression is elevated in human breast cancers and predicts poor survival in high-grade tumours. Finally, silencing of SCD in prostate orthografts efficiently blocked tumour growth and significantly increased animal survival. CONCLUSIONS: Our data implicate lipid desaturation as an essential process for cancer cell survival and suggest that targeting SCD could efficiently limit tumour expansion, especially under the metabolically compromised conditions of the tumour microenvironment.
RESUMO
Colorectal cancer (CRC) is one of the top three cancer-related causes of death worldwide. FBXW7 is a known tumor-suppressor gene, commonly mutated in CRC and in a variety of other epithelial tumors. Low expression of FBXW7 is also associated with poor prognosis. Loss of FBXW7 sensitizes cancer cells to certain drugs, while making them more resistant to other types of chemotherapies. However, is not fully understood how epithelial cells within normal gut and primary tumors respond to potential cancer therapeutics. We have studied genetically engineered mice in which the fbxw7 gene is conditionally knocked-out in the intestine (fbxw7(∆G)). To further investigate the mechanism of Fbxw7-action, we grew intestinal crypts from floxed-fbxw7 (fbxw7(fl/fl)) and fbxw7(ΔG) mice, in a Matrigel-based organoid (mini-gut) culture. The fbxw7(ΔG) organoids exhibited rapid budding events in the crypt region. Furthermore, to test organoids for drug response, we exposed day 3 intestinal organoids from fbxw7(fl/fl) and fbxw7(∆G) mice, to various concentrations of 5-fluorouracil (5-FU) for 72 hours. 5-FU triggers phenotypic differences in organoids including changing shape, survival, resistance, and death. 5-FU however, rescues the drug-resistance phenotype of fbxw7(ΔG) through the induction of terminal differentiation. Our results support the hypothesis that a differentiating therapy successfully targets FBXW7-mutated CRC cells.
RESUMO
The skin is a squamous epithelium that is continuously renewed by a population of basal layer stem/progenitor cells and can heal wounds. Here, we show that the transcription regulators YAP and TAZ localise to the nucleus in the basal layer of skin and are elevated upon wound healing. Skin-specific deletion of both YAP and TAZ in adult mice slows proliferation of basal layer cells, leads to hair loss and impairs regeneration after wounding. Contact with the basal extracellular matrix and consequent integrin-Src signalling is a key determinant of the nuclear localisation of YAP/TAZ in basal layer cells and in skin tumours. Contact with the basement membrane is lost in differentiating daughter cells, where YAP and TAZ become mostly cytoplasmic. In other types of squamous epithelia and squamous cell carcinomas, a similar control mechanism is present. By contrast, columnar epithelia differentiate an apical domain that recruits CRB3, Merlin (also known as NF2), KIBRA (also known as WWC1) and SAV1 to induce Hippo signalling and retain YAP/TAZ in the cytoplasm despite contact with the basal layer extracellular matrix. When columnar epithelial tumours lose their apical domain and become invasive, YAP/TAZ becomes nuclear and tumour growth becomes sensitive to the Src inhibitor Dasatinib.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Homeostase , Integrinas/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Fosfoproteínas/metabolismo , Transdução de Sinais , Pele/metabolismo , Animais , Proteínas de Ciclo Celular , Diferenciação Celular/efeitos dos fármacos , Diferenciação Celular/genética , Linhagem Celular , Núcleo Celular/efeitos dos fármacos , Núcleo Celular/metabolismo , Dasatinibe/farmacologia , Epitélio/efeitos dos fármacos , Epitélio/metabolismo , Receptores ErbB/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Homeostase/efeitos dos fármacos , Humanos , Queratinócitos/efeitos dos fármacos , Queratinócitos/metabolismo , Camundongos , Neoplasias de Células Escamosas/patologia , Fosfatidilinositol 3-Quinases/metabolismo , Estabilidade Proteica/efeitos dos fármacos , Transporte Proteico/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Pele/efeitos dos fármacos , Pele/patologia , Células-Tronco/citologia , Células-Tronco/efeitos dos fármacos , Células-Tronco/metabolismo , Transativadores , Fatores de Transcrição , Proteínas com Motivo de Ligação a PDZ com Coativador Transcricional , Cicatrização/efeitos dos fármacos , Proteínas de Sinalização YAP , Quinases da Família src/metabolismoRESUMO
In animals, the protein kinase C (PKC) family has expanded into diversely regulated subgroups, including the Rho family-responsive PKN kinases. Here, we describe knockouts of all three mouse PKN isoforms and reveal that PKN2 loss results in lethality at embryonic day 10 (E10), with associated cardiovascular and morphogenetic defects. The cardiovascular phenotype was not recapitulated by conditional deletion of PKN2 in endothelial cells or the developing heart. In contrast, inducible systemic deletion of PKN2 after E7 provoked collapse of the embryonic mesoderm. Furthermore, mouse embryonic fibroblasts, which arise from the embryonic mesoderm, depend on PKN2 for proliferation and motility. These cellular defects are reflected in vivo as dependence on PKN2 for mesoderm proliferation and neural crest migration. We conclude that failure of the mesoderm to expand in the absence of PKN2 compromises cardiovascular integrity and development, resulting in lethality.