Project description:The specific ablation of Rb1 gene in epidermis (RbF/F;K14cre) promotes proliferation and altered differentiation but does not produce spontaneous tumour development. These phenotypic changes are associated with increased expression of E2F members and E2F-dependent transcriptional activity. Here, we have focused on the possible dependence on E2F1 gene function. We have generated mice that lack Rb1 in epidermis in an inducible manner (RbF/F;K14creERTM). These mice are indistinguishable from those lacking pRb in this tissue in a constitutive manner (RbF/F;K14cre). In an E2F1-null background (RbF/F;K14creERTM; E2F1-/- mice), the phenotype due to acute Rb1 loss is not ameliorated by E2F1 loss, but rather exacerbated, indicating that pRb functions in epidermis do not rely solely on E2F1. On the other hand, RbF/F;K14creERTM;E2F1-/- mice develope spontaneous epidermal tumours of hair follicle origin with high incidence. These tumours, which retain a functional p19arf/p53 axis, also show aberrant activation of βcatenin/Wnt pathway. Gene expression studies revealed that these tumours display relevant similarities with specific human tumours. These data demonstrate that the Rb/E2F1 axis exerts essential functions not only in maintaining epidermal homeostasis, but also in suppressing tumour development in epidermis, and that the disruption of this pathway may induce tumour progression through specific alteration of developmental programs. Gene expression was compared between normal mouse skin, skin from transgenic RbF/F;K14creERTM; E2F1-/- , E2F1-/-, and RbF/F;K14creERTM; E2F1-/- mouse, and carcinomas arising in the skin of RbF/F;K14creERTM; E2F1-/- mouse. All mice were treated with tamoxifen.
Project description:The specific ablation of Rb1 gene in epidermis (RbF/F;K14cre) promotes proliferation and altered differentiation but does not produce spontaneous tumour development. These phenotypic changes are associated with increased expression of E2F members and E2F-dependent transcriptional activity. Here, we have focused on the possible dependence on E2F1 gene function. We have generated mice that lack Rb1 in epidermis in an inducible manner (RbF/F;K14creERTM). These mice are indistinguishable from those lacking pRb in this tissue in a constitutive manner (RbF/F;K14cre). In an E2F1-null background (RbF/F;K14creERTM; E2F1-/- mice), the phenotype due to acute Rb1 loss is not ameliorated by E2F1 loss, but rather exacerbated, indicating that pRb functions in epidermis do not rely solely on E2F1. On the other hand, RbF/F;K14creERTM;E2F1-/- mice develope spontaneous epidermal tumours of hair follicle origin with high incidence. These tumours, which retain a functional p19arf/p53 axis, also show aberrant activation of βcatenin/Wnt pathway. Gene expression studies revealed that these tumours display relevant similarities with specific human tumours. These data demonstrate that the Rb/E2F1 axis exerts essential functions not only in maintaining epidermal homeostasis, but also in suppressing tumour development in epidermis, and that the disruption of this pathway may induce tumour progression through specific alteration of developmental programs.
Project description:The specific deletion of Rb gene in epidermis leads to altered proliferation and differentiation, but not to the development of spontaneous tumors. Our previous data have demonstrated the existence of a functional compensation of Rb loss by Rbl1 (p107) in as the phenotypic differences with respect to controls are intensified. However, the possible evolution of this aggravated phenotype, in particular in relationship with tumorigenesis, has not been evaluated due to the premature death of the double deficient mice. We have now investigated whether p107 can also act as a tumor suppressor in pRb-deficient epidermis using different experimental approaches. We found spontaneous tumor development in doubly-deficient skin grafts. Moreover, Rb-deficient keratinocytes are susceptible to Ha-ras-induced transformation, and this susceptibility is enhanced by p107 loss. Further functional analyses, including microarray gene expression profiling, indicated that the loss of p107, in the absence of pRb, produces the reduction of p53-dependent pro-apoptotic signals. Overall, our data demonstrate that p107 behaves as a tumor suppressor in epidermis in the absence of pRb and suggest novel tumor-suppressive roles for p107 in the context of functional p53 and activated Ras Experiment Overall Design: Pools from RNA whole skin extracts from 3 animals of same genotype were done and analyzed, per duplicate, in mouse microarrays. Comparison was performed between the 4 different genotypes.
Project description:The specific deletion of Rb gene in epidermis leads to altered proliferation and differentiation, but not to the development of spontaneous tumors. Our previous data have demonstrated the existence of a functional compensation of Rb loss by Rbl1 (p107) in as the phenotypic differences with respect to controls are intensified. However, the possible evolution of this aggravated phenotype, in particular in relationship with tumorigenesis, has not been evaluated due to the premature death of the double deficient mice. We have now investigated whether p107 can also act as a tumor suppressor in pRb-deficient epidermis using different experimental approaches. We found spontaneous tumor development in doubly-deficient skin grafts. Moreover, Rb-deficient keratinocytes are susceptible to Ha-ras-induced transformation, and this susceptibility is enhanced by p107 loss. Further functional analyses, including microarray gene expression profiling, indicated that the loss of p107, in the absence of pRb, produces the reduction of p53-dependent pro-apoptotic signals. Overall, our data demonstrate that p107 behaves as a tumor suppressor in epidermis in the absence of pRb and suggest novel tumor-suppressive roles for p107 in the context of functional p53 and activated Ras Keywords: epidermis, tumorigenesis, pRb, p107, Ras, p53, apoptosis
Project description:The squamous cell carcinomas represent the aggressive type of non melanoma skin cancer, the most frequent malignancy among human population. We have studied here the possible relationship between these two pathways in skin using epidermal-specific mutant mice. Loss of p53, but not pRb, produces spontaneous tumor development, indicating that, contrary to pRb, p53 is the predominant tumor suppressor acting in mouse epidermis. The simultaneous inactivation of pRb and p53 does not aggravate the epidermal phenotype observed in Rb-deficient mice in terms of proliferation and/or differentiation. However, in doubly deficient mice spontaneous skin tumor development is severely accelerated. The tumors are aggressive, undifferentiated and display a hair follicle origin. Detailed analysis indicates that the acceleration is mediated by premature activation of the EGFR/Akt pathway, resulting in increased angiogenesis. The molecular characteristics of this model provide valuable tools to understand epidermal tumor formation, and may ultimately contribute to the development of therapies for the treatment of aggressive squamous cancer. Keywords: Squamous cell carcinoma, Epidermis, Tumorigenesis, pRb, p53, Akt, Angiogenesis
Project description:The squamous cell carcinomas represent the aggressive type of non melanoma skin cancer, the most frequent malignancy among human population. We have studied here the possible relationship between these two pathways in skin using epidermal-specific mutant mice. Loss of p53, but not pRb, produces spontaneous tumor development, indicating that, contrary to pRb, p53 is the predominant tumor suppressor acting in mouse epidermis. The simultaneous inactivation of pRb and p53 does not aggravate the epidermal phenotype observed in Rb-deficient mice in terms of proliferation and/or differentiation. However, in doubly deficient mice spontaneous skin tumor development is severely accelerated. The tumors are aggressive, undifferentiated and display a hair follicle origin. Detailed analysis indicates that the acceleration is mediated by premature activation of the EGFR/Akt pathway, resulting in increased angiogenesis. The molecular characteristics of this model provide valuable tools to understand epidermal tumor formation, and may ultimately contribute to the development of therapies for the treatment of aggressive squamous cancer. Experiment Overall Design: Pools from RNA whole skin extracts from 3 animals of same genotype were done and analyzed, per duplicate, in mouse microarrays. Comparison was performed between the 4 different genotypes.
Project description:The retinoblastoma protein (RB) is preferentially lost in the progression to castrate resistant prostate cancer (CRPC). However, the alterations associated with such loss have been scantly described. Current findings have identified a novel E2F1 associated cistrome and transcriptome that is associated with RB loss in PCa. In order to determine the contribution of chromatin accessibility to alterations in E2F1 activity, ATAC-Seq was performed.
Project description:E2F/RB activity is altered in most human tumors. The retinoblastoma family of proteins plays a key role in regulating the progression of the cell cycle from the G1 to S phases. This is achieved through negative regulation of E2F transcription factors, important positive regulators of cell cycle entry. E2F family members are divided in two groups: activators (E2F1-E2F3a) and repressors (E2F3b-E2F8). E2F4 accounts for a large part of the E2F activity and is a main E2F repressor member in vivo. Perturbations in the balance from quiescence towards proliferation contribute to increased mitotic gene expression levels frequently observed in cancer. We have previously reported that combined Rb1-Rbl1 and Rb1-E2F1 ablation in epidermis produces important alterations in epidermal proliferation and differentiation, leading to tumor development. However, the possible roles of E2F4 in this context are still to be determined. Here we show the absence of any discernible phenotype in the skin of mice lacking of E2F4. In contrast, the inducible loss of Rb1 in the epidermis of E2F4-null mice produced multiple skin abnormalities including altered differentiation and proliferation, spontaneous wounds, carcinoma in situ development and stem cell perturbations. All these phenotypic alterations are associated with extensive gene expression changes, the induction of c-myc and the Akt activation. Moreover, the whole transcriptome analyses in comparison with previous models generated also revealed extensive changes in multiple repressive complexes and in transcription factor activity. These results point to E2F4 as a master regulator in multiple steps of epidermal homeostasis in Rb1 absence.
Project description:The E2F family consists of transcriptional repressors and activators that control cell proliferation. In the classic paradigm of cell cycle regulation, the three activators, E2F1, E2F2 and E2F3, are invariably depicted as the final components of a CDK/Rb signaling cascade that executes the transcriptional program necessary to commit cells to enter S phase. Unexpectedly, we find through analysis of Affymetrix expression array data that mature lens epithelial cells deficient for E2F1-3 fail to repress cell cycle-regulated genes (and other targets of E2F) and that this corresponds with subsequent apoptosis and cellular collapse in the lens. Murine lenses were collected at two stages of development for RNA extraction and hybridization on Affymetrix microarrays. Our aim was to determine key events that lead to cellular collapse of lenses triply deficient for E2F1, E2F2, and E2F3 in neonates.