RESUMO
The contribution of deregulated chromatin architecture, including topologically associated domains (TADs), to cancer progression remains ambiguous. CCCTC-binding factor (CTCF) is a central regulator of higher-order chromatin structure that undergoes copy number loss in over half of all breast cancers, but the impact of this defect on epigenetic programming and chromatin architecture remains unclear. We find that under physiological conditions, CTCF organizes subTADs to limit the expression of oncogenic pathways, including phosphatidylinositol 3-kinase (PI3K) and cell adhesion networks. Loss of a single CTCF allele potentiates cell invasion through compromised chromatin insulation and a reorganization of chromatin architecture and histone programming that facilitates de novo promoter-enhancer contacts. However, this change in the higher-order chromatin landscape leads to a vulnerability to inhibitors of mTOR. These data support a model whereby subTAD reorganization drives both modification of histones at de novo enhancer-promoter contacts and transcriptional up-regulation of oncogenic transcriptional networks.
Assuntos
Montagem e Desmontagem da Cromatina , Regulação Neoplásica da Expressão Gênica , Invasividade Neoplásica , Fator de Ligação a CCCTC/metabolismo , Carcinogênese/genética , Cromatina/genética , Cromatina/metabolismo , Humanos , Invasividade Neoplásica/genética , Invasividade Neoplásica/patologia , Fosfatidilinositol 3-Quinases/genética , Fosfatidilinositol 3-Quinases/metabolismo , Regiões Promotoras GenéticasRESUMO
Epithelial tissues are highly organized structures that are structured at both the cellular and tissue levels. Individual cells are characterized by an apical membrane facing a central lumen, and a basolateral membrane that contacts adjacent cells and the basement membrane. The maintenance of apical-basal polarity is crucial for maintaining epithelial homeostasis and is considered a barrier to carcinogenesis. Apical-basal cell polarity is compromised in many epithelial cancers, such as breast, lung, and prostate, and has been associated with disease progression. Three-dimensional (3D) organotypic cultures recapitulate the 3D tissue architecture and mechanical properties found in vivo. This chapter describes methods to establish 3D organoids from human cell lines or mouse primary cells with inducible oncogene expression in polarized epithelial structures to investigate mechanisms of tumor initiation, luminal filling, and growth. The method is versatile, and simple modifications can be made to study diverse cell/tissue types and oncogenes.
Assuntos
Polaridade Celular , Células Epiteliais , Animais , Transformação Celular Neoplásica/metabolismo , Epitélio , Camundongos , OrganoidesRESUMO
Epithelial cell polarity defects support cancer progression. It is thus crucial to decipher the functional interactions within the polarity protein network. Here we show that Drosophila Girdin and its human ortholog (GIRDIN) sustain the function of crucial lateral polarity proteins by inhibiting the apical kinase aPKC. Loss of GIRDIN expression is also associated with overgrowth of disorganized cell cysts. Moreover, we observed cell dissemination from GIRDIN knockdown cysts and tumorspheres, thereby showing that GIRDIN supports the cohesion of multicellular epithelial structures. Consistent with these observations, alteration of GIRDIN expression is associated with poor overall survival in subtypes of breast and lung cancers. Overall, we discovered a core mechanism contributing to epithelial cell polarization from flies to humans. Our data also indicate that GIRDIN has the potential to impair the progression of epithelial cancers by preserving cell polarity and restricting cell dissemination.
Assuntos
Proteínas de Drosophila/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Proteínas dos Microfilamentos/metabolismo , Proteínas de Transporte Vesicular/metabolismo , Animais , Células CACO-2 , Diferenciação Celular/fisiologia , Polaridade Celular/fisiologia , Proliferação de Células/fisiologia , Proteínas de Drosophila/genética , Drosophila melanogaster , Células Epiteliais/citologia , Células Epiteliais/metabolismo , Feminino , Células HEK293 , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/genética , Masculino , Proteínas de Membrana/metabolismo , Proteínas dos Microfilamentos/genética , Morfogênese/fisiologia , Mapas de Interação de Proteínas , Proteína Quinase C/metabolismo , Proteínas de Transporte Vesicular/genéticaRESUMO
Breast cancer remains a leading cause of cancer-related death for women. The stepwise development of breast cancer through preinvasive to invasive disease is associated with progressive disruption of cellular and tissue organization. Apical-basal polarity is thought to be a barrier to breast cancer development, but the extent and potential mechanisms that contribute to disrupted polarity are incompletely understood. To investigate the cell polarity status of invasive breast cancers, we performed multiplex imaging of polarity markers on tissue cores from 432 patients from a spectrum of grades, stages and molecular subtypes. Apical-basal cell polarity was lost in 100% of cells in all cases studied, indicating that loss of epithelial polarity may be a universal feature of invasive breast cancer. We then analyzed genomic events from the TCGA dataset for an 18-gene set of core polarity genes. Coamplification of polarity genes with established breast oncogenes was found, which is consistent with functional cooperation within signaling amplicons. Gene-expression levels of several polarity genes were significantly associated with survival, and protein localization of Par6 correlated with higher grade, nodal metastasis and molecular subtype. Finally, multiple hotspot mutations in protein-protein interaction domains critical for cell polarity were identified. Our data indicate that genomic events likely contribute to pervasive disruption of epithelial polarity observed in invasive breast cancer.
