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Eran R Andrechek - One of the best experts on this subject based on the ideXlab platform.
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low E2F2 activity is associated with high genomic instability and parpi resistance
Scientific Reports, 2020Co-Authors: Jonathan P Rennhack, Eran R AndrechekAbstract:The E2F family, classically known for a central role in cell cycle, has a number of emerging roles in cancer including angiogenesis, metabolic reprogramming, metastasis and DNA repair. E2F1 specifically has been shown to be a critical mediator of DNA repair; however, little is known about DNA repair and other E2F family members. Here we present an integrative bioinformatic and high throughput drug screening study to define the role of E2F2 in maintaining genomic integrity in breast cancer. We utilized in vitro E2F2 ChIP-chip and over expression data to identify transcriptional targets of E2F2. This data was integrated with gene expression from E2F2 knockout tumors in an MMTV-Neu background. Finally, this data was compared to human datasets to identify conserved roles of E2F2 in human breast cancer through the TCGA breast cancer, Cancer Cell Line Encyclopedia, and CancerRx datasets. Through these methods we predict that E2F2 transcriptionally regulates mediators of DNA repair. Our gene expression data supports this hypothesis and low E2F2 activity is associated with a highly unstable tumor. In human breast cancer E2F2, status was also correlated with a patient's response to PARP inhibition therapy. Taken together this manuscript defines a novel role of E2F2 in cancer progression beyond cell cycle and could impact patient treatment.
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low E2F2 activity is associated with high genomic instability and parpi resistance
bioRxiv, 2019Co-Authors: Jonathan P Rennhack, Eran R AndrechekAbstract:Abstract The E2F family, classically known for a central role in cell cycle, has a number of emerging roles in cancer including angiogenesis, metabolic reprogramming, metastasis and DNA repair. E2F1 specifically has been shown to be a critical mediator of DNA repair; however, little is known about DNA repair and other E2F family members. Here we present an integrative bioinformatic and high throughput drug screening study to define the role of E2F2 in maintaining genomic integrity in breast cancer. We utilized in vitro E2F2 ChIP-chip and over expression data to identify transcriptional targets of E2F2. This data was integrated with gene expression from E2F2 knockout tumors in an MMTV-Neu background. Finally, this data was compared to human datasets to identify conserved roles of E2F2 in human breast cancer through the TCGA breast cancer, Cancer Cell Line Encyclopedia, and CancerRx datasets. Here we have computationally predicted that E2F2 transcriptionally regulates key mediators of DNA repair. Our gene expression data supports this hypothesis and low E2F2 activity is associated with a highly unstable tumor. In human breast cancer E2F2, status was also correlated with a patient’s response to PARP inhibition therapy. Taken together this manuscript defines a novel role of E2F2 in cancer progression beyond cell cycle and could be therapeutically relevant. Author Summary The E2F family of proteins have been known to regulate cell cycle and have recently been shown to have a number of roles in tumor progression. Here we use a combination of computational techniques and high-throughput drug screening data to establish a novel role of E2F2 in maintaining genomic integrity. We have shown that a number of direct and indirect target genes of E2F2 are involved in multiple classical DNA repair pathways. Importantly, this was shown to be unique to E2F2 and not present with other activator E2Fs like E2F1. We have also shown that E2F2 activity is positively correlated with PARP inhibitor sensitivity regardless of BRCA1/2 status. This is important due to the recent approval of PARP inhibitor therapy in the clinic. Based on our work E2F2 activity could serve as a novel biomarker of response and may identify a new cohort of patients which could benefit from PARPi therapy.
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transcription factor compensation during mammary gland development in e2f knockout mice
PLOS ONE, 2018Co-Authors: Eran R AndrechekAbstract:The E2F transcription factors control key elements of development, including mammary gland branching morphogenesis, with several E2Fs playing essential roles. Additional prior data has demonstrated that loss of individual E2Fs can be compensated by other E2F family members, but this has not been tested in a mammary gland developmental context. Here we have explored the role of the E2Fs and their ability to functionally compensate for each other during mammary gland development. Using gene expression from terminal end buds and chromatin immunoprecipitation data for E2F1, E2F2 and E2F3, we noted both overlapping and unique mammary development genes regulated by each of the E2Fs. Based on our computational findings and the fact that E2Fs share a common binding motif, we hypothesized that E2F transcription factors would compensate for each other during mammary development and function. To test this hypothesis, we generated RNA from E2F1-/-, E2F2-/- and E2F3+/- mouse mammary glands. QRT-PCR on mammary glands during pregnancy demonstrated increases in E2F2 and E2F3a in the E2F1-/- mice and an increase in E2F2 levels in E2F3+/- mice. During lactation we noted that E2F3b transcript levels were increased in the E2F2-/- mice. Given that E2Fs have previously been noted to have the most striking effects on development during puberty, we hypothesized that loss of individual E2Fs would be compensated for at that time. Double mutant mice were generated and compared with the single knockouts. Loss of both E2F1 and E2F2 revealed a more striking phenotype than either knockout alone, indicating that E2F2 was compensating for E2F1 loss. Interestingly, while E2F2 was not able to functionally compensate for E2F3+/- during mammary outgrowth, increased E2F2 expression was observed in E2F3+/- mammary glands during pregnancy day 14.5 and lactation day 5. Together, these findings illustrate the specificity of E2F family members to compensate during development of the mammary gland.
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abstract a79 elucidating the role of E2F2 transcription factor in mediating human breast cancer metastasis
Cancer Research, 2013Co-Authors: Inez Yuwanita, Danielle Barnes, Eran R AndrechekAbstract:In human breast cancer metastasis, it is not the primary tumor that is the cause of mortality but metastasis to distant sites. Therefore, it is important to elucidate the biological mechanism that underlies the metastatic processes. To address this, we utilized bioinformatic methods in combination with biochemical assays and genetic studies. Specifically, we crossed a MMTV-Myc mouse model with various activator E2Fs mutants and found that the tumor incidence decreased in the MMTV-Myc mice crossed with E2F2 and E2F3 mutant background. Surprisingly, we also found that the loss of E2F2 sharply increased lung metastasis in MMTV-Myc mice. Here, we test whether these observations in mice translate to human breast cancer and identify the genes under the control of E2F2 transcription factor that are responsible for mediating breast cancer metastasis. To achieve these aims, we examined the probability of activation of the E2F2 transcription factors in human breast cancer. We calculated the probability of E2F2 activation and used this data to stratify the human breast cancer samples and their associated metastatic clinical data. We performed hierarchical clustering of human and mouse gene expression datasets and uncovered two clusters of human breast tumors in which E2F2 played distinctly different roles in mediating metastasis. The human breast cancer samples that clustered with our mouse gene expression data exhibited the same role for E2F2 whereby it increased time to distant metastasis. We then examined genes that were differentially regulated within this cluster and found that there was a set of genes that were associated with a difference in human metastasis survival times and with the E2F2 knockout in the mouse model. To further explore the mechanism by which E2F2 mediates human breast cancer metastasis, we have identified four genes asputative E2F2 target genes including KLK1, MYH2, PTPRD and TNNC2. We found that partial knockdown of KLK1 promotes the migratory capacity of MCF7 cell in vitro. In addition, partial knockdown of PTPRD affected cellular morphology, where they were found to grow in discreet clusters while having little effect on the migration capacity in a scratch assay. We are currently exploring the effects of MYH2 and TNNC2 ablation on these cells. Taken together, we have begun to elucidate the roles of E2F2 transcription factor in mediating breast cancer metastasis as well as establishing a mechanism by which E2F2 regulates metastasis. Citation Format: Inez Yuwanita, Danielle Barnes, Eran Andrechek. Elucidating the role of E2F2 transcription factor in mediating human breast cancer metastasis. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Invasion and Metastasis; Jan 20-23, 2013; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2013;73(3 Suppl):Abstract nr A79.
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abstract 1286 elucidating the role of E2F2 in mediating human breast cancer metastasis
Cancer Research, 2012Co-Authors: Inez Yuwanita, Danielle Barnes, Eran R AndrechekAbstract:Proceedings: AACR 103rd Annual Meeting 2012‐‐ Mar 31‐Apr 4, 2012; Chicago, IL In human breast cancer it is not the primary tumor that is the cause of mortality but tumor metastasis to distant sites. Therefore, it is important to elucidate the biological mechanisms that underlie the metastatic processes. To address this, we utilized bioinformatic methods in combination with biochemical assays and genetic studies. Specifically, we crossed MMTV-Myc transgenic mice with various activator E2Fs mutants. We found that the tumor incidence decreased in MMTV-Myc mice interbred with E2F2 and E2F3 mutant backgrounds. Surprisingly, we also found that the loss of E2F2 sharply increased the percentage of lung metastasis in MMTV-Myc mice. Here, we test whether these observations in mice translate to human breast cancer and identify the genes under the control of E2F2 transcription factor that are responsible for mediating breast cancer metastasis. To achieve these aims, we examined the probability of activation of the E2F2 transcription factors in human breast cancer. We calculated the probability of E2F2 activation and used this data to stratify the human breast cancer samples and their associated metastatic clinical data. Examining this data we found that a low probability of E2F2 pathway activation led to decreased time to distant metastasis. To account for this discrepancy, we performed hierarchical clustering of human and mouse gene expression datasets. We found two clusters of human breast tumors in which E2F2 played distinctly different roles in mediating metastasis. One particular cluster clustered with our mouse gene expression data and exhibited the same role for E2F2 where it increased time to distant metastasis. In addition, we examined the ontology of the genes in this cluster and found that the majority of these genes had putative E2F binding sites. To further identify genes that are involved in metastasis, we stratified human breast cancer metastasis clinical annotation using gene expression values from these genes. We found that there were seven genes that are uniquely differentially regulated in this cluster. Interestingly, we observed that this group of seven genes included genes that were implicated in the metastatic process such as MMP16. Taken together, our results showed that there is an important role for E2F2 in mediating breast cancer metastasis and using our bioinformatic methods, we have begun to elucidate this role of E2F2 in mediating breast cancer metastasis. Our future directions include further validation of these target genes in vitro and in vivo to establish the mechanism by which E2F2 mediates breast cancer metastasis. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1286. doi:1538-7445.AM2012-1286
Joseph R Nevins - One of the best experts on this subject based on the ideXlab platform.
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interaction of e2f7 transcription factor with e2f1 and c terminal binding protein ctbp provides a mechanism for e2f7 dependent transcription repression
Journal of Biological Chemistry, 2013Co-Authors: Beiyu Liu, Igor Shats, Steven P Angus, Michael L Gatza, Joseph R NevinsAbstract:Previous work has identified distinct functions for E2F proteins during a cellular proliferative response including a role for E2F1–3 in the activation of transcription at G1/S and a role for E2F4–8 in repressing the same group of E2F1–3 target genes as cells progress through S phase. We now find that E2F7 and E2F8, which are induced by E2F1–3 at G1/S, can form a heterodimer with E2F1 through interactions involving the DNA-binding domains of the two proteins. In vitro DNA interaction assays demonstrate the formation of an E2F1-E2F7 complex, as well as an E2F7-E2F7 complex on adjacent E2F-binding sites. We also show that E2F7 recruits the co-repressor C-terminal-binding protein (CtBP) and that CtBP2 is essential for E2F7 to repress E2F1 transcription. Taken together, these findings suggest a mechanism for the repression of transcription by E2F7. Background: E2F7 is a transcription factor that controls cell cycle by repressing the expression of G1/S genes in late S phase. Results: E2F7 forms a heterodimer with E2F1, and it recruits the co-repressor CtBP to repress G1/S transcription. Conclusion: E2F7 represses gene transcription by interacting with E2F1 and co-repressor CtBP. Significance: These findings suggest a mechanism for the repression of transcription by E2F7.
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specificity in the activation and control of transcription factor e2f dependent apoptosis
Proceedings of the National Academy of Sciences of the United States of America, 2003Co-Authors: Timothy C Hallstrom, Joseph R NevinsAbstract:Previous work has demonstrated a role for the E2F1 gene product in signaling apoptosis, both as a result of the deregulation of the Rb/E2F pathway as well as in response to DNA damage. We now show that the ability of cells to suppress the apoptotic potential of E2F1, as might occur during the course of normal cellular proliferation, requires the action of the Ras–phosphoinositide 3-kinase–Akt signaling pathway. In addition, we also identify a domain within the E2F1 protein, previously termed the marked-box domain, that is essential for the apoptotic activity of E2F1 and that distinguishes the E2F1 protein from E2F3. We also show that the E2F1-marked-box domain is essential for the induction of both p53 and p73 accumulation. Importantly, a role for the marked-box domain in the specificity of E2F1-mediated apoptosis coincides with recent work demonstrating a role for this domain in achieving specificity in the activation of transcription. We conclude that the unique capacity of E2F1 to trigger apoptosis reflects a specificity of transcriptional activation potential, and that this role for E2F1 is regulated through the action of the Akt protein kinase.
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identification of e box factor tfe3 as a functional partner for the e2f3 transcription factor
Molecular and Cellular Biology, 2003Co-Authors: Paloma H Giangrande, Timothy C Hallstrom, Chainarong Tunyaplin, Kathryn Calame, Joseph R NevinsAbstract:The ability of the retinoblastoma (Rb) tumor suppressor protein to regulate cell growth is due, at least in part, to its ability to interact with and regulate the E2F family of transcription factors (8, 34). The E2F proteins have been shown to control the expression of a large number of genes involved in DNA replication, cell cycle progression, and cell fate determination. The E2F family is composed of six distinct gene products that form heterodimeric complexes with partners of the DP family. Sequence analysis reveals three distinct subfamilies of E2F genes: the E2F1, E2F2, and E2F3 genes, the E2F4 and E2F5 genes, and the E2F6 gene. This division also coincides with functional distinctions. The E2F1, E2F2, and E2F3 genes are tightly regulated by cell growth and during the cell cycle, whereas E2F4, E2F5 and E2F6 are constitutively expressed. This cell cycle regulation of E2F1, E2F2, and E2F3 transcription is complemented by mechanisms that tightly regulate the accumulation of the proteins. An N-terminal domain unique to E2F1 to E2F3 is responsible for both ubiquitin-mediated degradation of the proteins (29) and targeting by the cyclin A/cdk2 kinase, the latter leading to inhibition of DNA binding capacity (7, 22, 23, 53). The E2F proteins also vary in their role as transcriptional regulatory activities. While E2F1 to E2F3 act as positive regulators of transcription, E2F4 and E2F5 appear to function primarily as transcriptional repressors in concert with Rb family members. E2F6 also appears to function as a transcriptional repressor but in a manner independent of Rb (4, 10, 47, 48). Various experiments have suggested distinct functional roles for the activating E2F proteins E2F1, E2F2, and E2F3. The E2F3 protein appears to be particularly important for cell proliferation, as seen from the inhibition of E2F3 activity by antibody microinjection (25) as well as the results of deletion of the E2F3 gene (18). Moreover, the expression of a number of E2F target genes that encode key cell cycle regulatory proteins, including B-Myb, cyclin A, cdc2, cdc6, and dihydrofolate reductase, are reduced in E2F3 null fibroblasts but not in E2F1 null cells (18). In contrast, E2F1 appears to play a role in triggering an apoptotic response, either when overexpressed in the absence of survival signals (6, 21, 39, 43, 51) or in response to DNA damage (27). In addition, the ability of Myc to induce apoptosis is impaired in the absence of E2F1 function but unaltered by the absence of either E2F2 or E2F3 (26). Given the role of the E2F1, E2F2, and E2F3 proteins as transcriptional activators, the specificity of function might best be explained by an ability of these E2F proteins to activate distinct target genes that then carry out these functions of apoptosis or proliferation. As an example, the p19ARF gene has been shown to be an E2F1 target gene (2, 6), linking the action of the Rb/E2F pathway with the p53 response leading to apoptosis. Similarly, the Apaf1 gene appears to be activated specifically by E2F1 (30). Although one potential mechanism for such specificity could be an ability of these proteins to recognize subtle differences in cis-acting promoter sequences, there is little evidence for distinct DNA sequence recognition among the E2F isoforms. More importantly, analysis of the structure of an E2F-DNA complex did not show a capacity for distinct DNA sequence recognition when the amino acid variation within the E2F family is considered (56). An alternative mechanism for promoter specificity could involve distinct protein-protein interactions. Possibly, sequences within E2F3 allow interaction with a subset of cellular proteins that provide a basis for promoter specificity and that are distinct from the proteins that can interact with E2F1. For example, the interaction of the herpesvirus VP16 transcriptional activator protein with the cellular factor HCF-1 and the cellular Oct-1 transcription factor redirects Oct-1 to herpesvirus immediate-early promoters by virtue of expanded DNA sequence recognition (1). A further example of combinatorial specificity as a mechanism for the specificity of transcription factor function is the pancreatic islet factor STF-1, which interacts with Pbx in a cooperative fashion and targets Pbx to a subset of promoters containing STF-1 binding sites (38). Recently, we identified the YY1-binding protein RYBP as a factor which binds to E2F2 and E2F3 but not E2F1 and recruits these E2Fs to a subset of E2F target promoters containing YY1 binding sites (41). To further explore the mechanistic basis for the specificity of E2F transcription activation, we used yeast two-hybrid screens to identify proteins that specifically interact with E2F3. In so doing, we identified the TFE3 transcription factor as a specific partner for E2F3. Recent experiments identified TFE3 as an activity that can rescue Rb-mediated growth arrest, providing a functional link between TFE3 and the Rb/E2F pathway (M. Nijman, S. Hijmans, and R. Bernards, personal communication). Moreover, previous work identified TFE3 as a fusion partner in chromosomal rearrangements in renal cell carcinomas (14, 45, 50). We further show that TFE3 and E2F3 can act synergistically to activate the p68 subunit gene of DNA polymerase α, dependent on the ability of the two proteins to physically interact, and that the two activities associate with the p68 promoter within intact cells in a mutually dependent manner.
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gene expression phenotypic models that predict the activity of oncogenic pathways
Nature Genetics, 2003Co-Authors: Erich Huang, Seiichi Ishida, Jennifer Pittman, Holly K Dressman, Andrea Bild, Mark T Kloos, Mark Damico, Richard G Pestell, Michael West, Joseph R NevinsAbstract:High-density DNA microarrays measure expression of large numbers of genes in one assay. The ability to find underlying structure in complex gene expression data sets and rigorously test association of that structure with biological conditions is essential to developing multi-faceted views of the gene activity that defines cellular phenotype. We sought to connect features of gene expression data with biological hypotheses by integrating 'metagene' patterns from DNA microarray experiments in the characterization and prediction of oncogenic phenotypes. We applied these techniques to the analysis of regulatory pathways controlled by the genes HRAS (Harvey rat sarcoma viral oncogene homolog), MYC (myelocytomatosis viral oncogene homolog) and E2F1, E2F2 and E2F3 (encoding E2F transcription factors 1, 2 and 3, respectively). The phenotypic models accurately predict the activity of these pathways in the context of normal cell proliferation. Moreover, the metagene models trained with gene expression patterns evoked by ectopic production of Myc or Ras proteins in primary tissue culture cells properly predict the activity of in vivo tumor models that result from deregulation of the MYC or HRAS pathways. We conclude that these gene expression phenotypes have the potential to characterize the complex genetic alterations that typify the neoplastic state, whether in vitro or in vivo, in a way that truly reflects the complexity of the regulatory pathways that are affected.
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interaction of yy1 with e2fs mediated by rybp provides a mechanism for specificity of e2f function
The EMBO Journal, 2002Co-Authors: Susanne Schlisio, Joseph R Nevins, Terri J Halperin, Miguel VidalAbstract:To explore mechanisms for specificity of function within the family of E2F transcription factors, we have identified proteins that interact with individual E2F proteins. A two-hybrid screen identified RYBP (Ring1- and YY1-binding protein) as a protein that interacts specifically with the E2F2 and E2F3 family members, dependent on the marked box domain in these proteins. The Cdc6 promoter contains adjacent E2F- and YY1-binding sites, and both are required for promoter activity. In addition, YY1 and RYBP, in combination with either E2F2 or E2F3, can stimulate Cdc6 promoter activity synergistically, dependent on the marked box domain of E2F3. Using chromatin immunoprecipitation assays, we show that both E2F2 and E2F3, as well as YY1 and RYBP, associate with the Cdc6 promoter at G1/S of the cell cycle. In contrast, we detect no interaction of E2F1 with the Cdc6 promoter. We suggest that the ability of RYBP to mediate an interaction between E2F2 or E2F3 and YY1 is an important component of Cdc6 activation and provides a basis for specificity of E2F function.
Gustavo Leone - One of the best experts on this subject based on the ideXlab platform.
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division and apoptosis of e2f deficient retinal progenitors
Nature, 2009Co-Authors: Danian Chen, Gustavo Leone, Marek Pacal, Pamela L Wenzel, Paul S Knoepfler, Rod BremnerAbstract:The activating E2f transcription factors (E2f1, E2F2 and E2f3) induce transcription and are widely viewed as essential positive cell cycle regulators. Indeed, they drive cells out of quiescence, and the 'cancer cell cycle' in Rb1 null cells is E2f-dependent. Absence of activating E2fs in flies or mammalian fibroblasts causes cell cycle arrest, but this block is alleviated by removing repressive E2f or the tumour suppressor p53, respectively. Thus, whether activating E2fs are indispensable for normal division is an area of debate. Activating E2fs are also well known pro-apoptotic factors, providing a defence against oncogenesis, yet E2f1 can limit irradiation-induced apoptosis. In flies this occurs through repression of hid (also called Wrinkled; Smac/Diablo in mammals). However, in mammals the mechanism is unclear because Smac/Diablo is induced, not repressed, by E2f1, and in keratinocytes survival is promoted indirectly through induction of DNA repair targets. Thus, a direct pro-survival function for E2f1-3 and/or its relevance beyond irradiation has not been established. To address E2f1-3 function in normal cells in vivo we focused on the mouse retina, which is a relatively simple central nervous system component that can be manipulated genetically without compromising viability and has provided considerable insight into development and cancer. Here we show that unlike fibroblasts, E2f1-3 null retinal progenitor cells or activated Muller glia can divide. We attribute this effect to functional interchangeability with Mycn. However, loss of activating E2fs caused downregulation of the p53 deacetylase Sirt1, p53 hyperacetylation and elevated apoptosis, establishing a novel E2f-Sirt1-p53 survival axis in vivo. Thus, activating E2fs are not universally required for normal mammalian cell division, but have an unexpected pro-survival role in development.
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intestinal hyperplasia induced by simian virus 40 large tumor antigen requires E2F2
Journal of Virology, 2007Co-Authors: Teresa M Saenzrobles, Rene Opavsky, Gustavo Leone, Jennifer A Markovics, Jean Leon Chong, Robert H Whitehead, James M PipasAbstract:The simian virus 40 large T antigen contributes to neoplastic transformation, in part, by targeting the Rb family of tumor suppressors. There are three known Rb proteins, pRb, p130, and p107, all of which block the cell cycle by preventing the transcription of genes regulated by the E2F family of transcription factors. T antigen interacts directly with Rb proteins and disrupts Rb-E2F complexes both in vitro and in cultured cells. Consequently, T antigen is thought to inhibit transcriptional repression by the Rb family proteins by disrupting their interaction with E2F proteins, thus allowing E2F-dependent transcription and the expression of cellular genes needed for entry into S phase. This model predicts that active E2F-dependent transcription is required for T-antigen-induced transformation. To test this hypothesis, we have examined the status of Rb-E2F complexes in murine enterocytes. Previous studies have shown that T antigen drives enterocytes into S phase, resulting in intestinal hyperplasia, and that the induction of enterocyte proliferation requires T-antigen binding to Rb proteins. In this paper, we show that normal growth-arrested enterocytes contain p130-E2F4 complexes and that T-antigen expression destroys these complexes, most likely by stimulating p130 degradation. Furthermore, unlike their normal counterparts, enterocytes expressing T antigen contain abundant levels of E2F2 and E2F3a. Concomitantly, T-antigen-induced intestinal proliferation is reduced in mice lacking either E2F2 alone or both E2F2 and E2F3a, but not in mice lacking E2F1. These studies support a model in which T antigen eliminates Rb-E2F repressive complexes so that specific activator E2Fs can drive S-phase entry.
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e2f1 E2F2 and e2f3 control e2f target expression and cellular proliferation via a p53 dependent negative feedback loop
Molecular and Cellular Biology, 2007Co-Authors: Cynthia Timmers, Baidehi Maiti, Rene Opavsky, Harold I Saavedra, Nidhi Sharma, Daniel A Orringer, Prashant Trikha, Gustavo LeoneAbstract:E2F-mediated control of gene expression is believed to have an essential role in the control of cellular proliferation. Using a conditional gene-targeting approach, we show that the targeted disruption of the entire E2F activator subclass composed of E2f1, E2F2, and E2f3 in mouse embryonic fibroblasts leads to the activation of p53 and the induction of p53 target genes, including p21 CIP1 . Consequently, cyclin-dependent kinase activity and retinoblastoma (Rb) phosphorylation are dramatically inhibited, leading to Rb/E2F-mediated repression of E2F target gene expression and a severe block in cellular proliferation. Inactivation of p53 in E2f1-, E2F2-, and E2f3-deficient cells, either by spontaneous mutation or by conditional gene ablation, prevented the induction of p21 CIP1 and many other p53 target genes. As a result, cyclin-dependent kinase activity, Rb phosphorylation, and E2F target gene expression were restored to nearly normal levels, rendering cells responsive to normal growth signals. These findings suggest that a critical function of the E2F1, E2F2, and E2F3 activators is in the control of a p53-dependent axis that indirectly regulates E2F-mediated transcriptional repression and cellular proliferation.
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control of the p53 p21cip1 axis by e2f1 E2F2 and e2f3 is essential for g1 s progression and cellular transformation
Journal of Biological Chemistry, 2006Co-Authors: Nidhi Sharma, Cynthia Timmers, Harold I Saavedra, Prashant Trikha, Amanda Obery, Gustavo LeoneAbstract:The E2F family of transcription factors is believed to have an essential role in the control of cellular proliferation by regulating the transcription of genes involved in cell cycle progression. Previous work has demonstrated that the targeted inactivation of E2f1, E2F2, and E2f3 results in elevated p21CIP1 protein levels, loss of E2F target gene expression, and cell cycle arrest at G1/S and G2/M, suggesting a strict requirement for these E2Fs in the control of normal cellular proliferation. We now demonstrate that E2f1, E2F2, and E2f3 are also required for oncogene-mediated transformation of mouse embryonic fibroblasts. Analysis of synchronized populations of mouse embryonic fibroblasts revealed that the inactivation of p21CIP1 restores the ability of E2f1-3-deficient cells to enter and transit through G1/S (but not G2/M). In contrast, loss of p53 restored the ability of these cells to progress through both G1/S and mitosis, leading to their continued proliferation. The inactivation of p53 (but not p21CIP1) rendered E2f1-3-deficient cells sensitive to transformation and tumorigenesis. These results suggest that the negative regulation of the p53-p21CIP1 axis by the E2F1-3 factors is critical for cell cycle progression and cellular transformation.
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cloning and characterization of mouse e2f8 a novel mammalian e2f family member capable of blocking cellular proliferation
Journal of Biological Chemistry, 2005Co-Authors: Baidehi Maiti, Alain De Bruin, Faye Gordon, Cynthia Timmers, Rene Opavsky, Kaustubha Patil, John Tuttle, Whitney M Cleghorn, Gustavo LeoneAbstract:The E2F transcription factor family plays a crucial and well established role in cell cycle progression. Deregulation of E2F activities in vivo leads to developmental defects and cancer. Based on current evidence in the field, mammalian E2Fs can be functionally categorized into either transcriptional activators (E2F1, E2F2, and E2F3a) or repressors (E2F3b, E2F4, E2F5, E2F6, and E2F7). We have identified a novel E2F family member, E2F8, which is conserved in mice and humans and has its counterpart in Arabidopsis thaliana (E2Ls). Interestingly, E2F7 and E2F8 share unique structural features that distinguish them from other mammalian E2F repressor members, including the presence of two distinct DNA-binding domains and the absence of DP-dimerization, retinoblastoma-binding, and transcriptional activation domains. Similar to E2F7, overexpression of E2F8 significantly slows down the proliferation of primary mouse embryonic fibroblasts. These observations, together with the fact that E2F7 and E2F8 can homodimerize and are expressed in the same adult tissues, suggest that they may have overlapping and perhaps synergistic roles in the control of cellular proliferation.
James Degregori - One of the best experts on this subject based on the ideXlab platform.
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e2f1 and E2F2 are differentially required for homeostasis driven and antigen induced t cell proliferation in vivo
Journal of Immunology, 2005Co-Authors: Deborah Deryckere, James DegregoriAbstract:Homeostasis-driven T cell proliferation occurs in response to a lymphopenic environment and is mediated by TCR and IL-7 signaling. In this report, we demonstrate a defect in the proliferation of murine naive and memory T cells lacking both E2F1 and E2F2 in response to lymphopenic conditions, suggesting that E2F1 and E2F2 function redundantly downstream of TCR and/or IL-7 signaling during homeostasis-driven proliferation. In contrast, T cell proliferation in response to antigenic stimulation is either unaffected (in vivo) or potentiated (ex vivo) by loss of E2F1 and E2F2, indicating divergent requirements for these E2F factors in T cell proliferation mediated by distinct stimuli. E2F1/E2F2 double knockout (DKO) T cells enter S phase in response to homeostatic signaling, but fail to divide, suggesting that S phase progression is either incomplete or defective. In addition, E2F1/E2F2 DKO mice do not recover normal T cell numbers following exposure to a sublethal dose of radiation, indicating that this defect in homeostasis-driven proliferation is physiologically relevant. Consistent with their failure in cell cycle progression, the differentiation of DKO T cells into memory T cells in response to homeostatic signals is significantly reduced. These observations support the idea that proliferation is required for memory T cell formation and also have implications for the development of clinical strategies to minimize the occurrence of lymphopenia-induced autoimmunity.
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the development of diabetes in e2f1 E2F2 mutant mice reveals important roles for bone marrow derived cells in preventing islet cell loss
Proceedings of the National Academy of Sciences of the United States of America, 2003Co-Authors: Jing W Zhu, Christopher J Hogan, Jeffery S Tessem, Joshua Beilke, Marileila Varellagarcia, Jan Jensen, James DegregoriAbstract:Our studies of mice deficient for the E2F1 and E2F2 transcription factors have revealed essential roles for these proteins in the cell cycle control of pancreatic exocrine cells and the regulation of pancreatic beta cell maintenance. Pancreatic exocrine cells in E2f1/E2F2 mutant mice become increasingly polyploid with age, coinciding with severe exocrine atrophy. Furthermore, mice deficient for both E2F1 and E2F2 develop nonautoimmune, insulin-dependent diabetes with high penetrance. Surprisingly, transplantation of wild-type bone marrow can prevent or rescue diabetes in E2f1-/-E2F2-/- mice. We hypothesize that exocrine degeneration results in a destructive environment for beta cells, which can be alleviated by restoration of the hematopoietic system that is also defective in E2f1-/-E2F2-/- mice. The demonstration that beta cell maintenance under conditions of stress is influenced by bone marrow-derived cells may provide important insight into the design of therapies to boost islet mass and function in diabetic patients.
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defective gene expression s phase progression and maturation during hematopoiesis in e2f1 E2F2 mutant mice
Molecular and Cellular Biology, 2003Co-Authors: Jing W Zhu, Christopher J Hogan, James DegregoriAbstract:E2F plays critical roles in cell cycle progression by regulating the expression of genes involved in nucleotide synthesis, DNA replication, and cell cycle control. We show that the combined loss of E2F1 and E2F2 in mice leads to profound cell-autonomous defects in the hematopoietic development of multiple cell lineages. E2F2 mutant mice show erythroid maturation defects that are comparable with those observed in patients with megaloblastic anemia. Importantly, hematopoietic defects observed in E2F1/E2F2 double-knockout (DKO) mice appear to result from impeded S phase progression in hematopoietic progenitor cells. During DKO B-cell maturation, differentiation beyond the large pre-BII-cell stage is defective, presumably due to failed cell cycle exit, and the cells undergo apoptosis. However, apoptosis appears to be the consequence of failed maturation, not the cause. Despite the accumulation of hematopoietic progenitor cells in S phase, the combined loss of E2F1 and E2F2 results in significantly decreased expression and activities of several E2F target genes including cyclin A2. Our results indicate specific roles for E2F1 and E2F2 in the induction of E2F target genes, which contribute to efficient expansion and maturation of hematopoietic progenitor cells. Thus, E2F1 and E2F2 play essential and redundant roles in the proper coordination of cell cycle progression with differentiation which is necessary for efficient hematopoiesis.
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distinct roles for e2f proteins in cell growth control and apoptosis
Proceedings of the National Academy of Sciences of the United States of America, 1997Co-Authors: James Degregori, Gustavo Leone, Alexander Miron, Laszlo Jakoi, Joseph R NevinsAbstract:E2F transcription activity is composed of a family of heterodimers encoded by distinct genes. Through the overproduction of each of the five known E2F proteins in mammalian cells, we demonstrate that a large number of genes encoding proteins important for cell cycle regulation and DNA replication can be activated by the E2F proteins and that there are distinct specificities in the activation of these genes by individual E2F family members. Coexpression of each E2F protein with the DP1 heterodimeric partner does not significantly alter this specificity. We also find that only E2F1 overexpression induces cells to undergo apoptosis, despite the fact that at least two other E2F family members, E2F2 and E2F3, are equally capable of inducing S phase. The ability of E2F1 to induce apoptosis appears to result from the specific induction of an apoptosis-promoting activity rather than the lack of induction of a survival activity, because co-expression of E2F2 and E2F3 does not rescue cells from E2F1-mediated apoptosis. We conclude that E2F family members play distinct roles in cell cycle control and that E2F1 may function as a specific signal for the initiation of an apoptosis pathway that must normally be blocked for a productive proliferation event.
Akihiko Tsuji - One of the best experts on this subject based on the ideXlab platform.
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transcriptional regulation of fibrillin 2 gene by e2f family members in chondrocyte differentiation
Journal of Cellular Biochemistry, 2009Co-Authors: Takuya Yanagino, Keizo Yuasa, Yoshiko Matsuda, Masami Nagahama, Akihiko TsujiAbstract:Mutation in fibrillin-2, a major structural component of extracellular microfibrils in connective tissue, results in the autosomal dominant disease congenital contractural arachnodactyly. This genetic disease is characterized by dolichostenomelia and arachnodactyly, in addition to contractures of the large joints and abnormal pinnae formation, thus indicating the significance of fibrillin-2 in chondrogenesis. In this study, we investigated the transcriptional regulation of fibrillin-2 in chondrogenic differentiation. Although mRNA expression of fibrillin-1, a highly homologous protein to fibrillin-2, remained almost unchanged during chondrogenesis of mouse ATDC5 cells, fibrillin-2 mRNA expression varied. Fibrillin-2 was highly expressed at the early stage and declined progressively during differentiation. The 5′-flanking region of the fibrillin-2 gene contains potential binding sites for E2F, Runx, AP-2, and Sox transcription factors. The promoter activity of fibrillin-2 decreased markedly following deletion and mutagenesis of the E2F binding site between −143 and −136 bp. Overexpression of E2F1 resulted in a marked increase in its promoter activity, whereas expression of other transcription factors including AP-2α and Runx2 had no effect. The increase in promoter activity by E2F1 was completely suppressed by the coexpression of E2F4. E2F2 and E2F3 had positive effects on the promoter activity. Although ATDC5 cells expressed transcripts for the E2F family genes at all stages of differentiation, the expression profiles differed. E2F1 expression remained almost unchanged, whereas E2F4 expression increased markedly at the late stage of differentiation. These results indicated that coordinated expression of the E2F family is critical for the transcriptional regulation of fibrillin-2 during chondrogenesis. J. Cell. Biochem. 106: 580–588, 2009. © 2009 Wiley-Liss, Inc.
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transcriptional regulation of fibrillin 2 gene by e2f family members in chondrocyte differentiation
Journal of Cellular Biochemistry, 2009Co-Authors: Takuya Yanagino, Keizo Yuasa, Yoshiko Matsuda, Masami Nagahama, Akihiko TsujiAbstract:Mutation in fibrillin-2, a major structural component of extracellular microfibrils in connective tissue, results in the autosomal dominant disease congenital contractural arachnodactyly. This genetic disease is characterized by dolichostenomelia and arachnodactyly, in addition to contractures of the large joints and abnormal pinnae formation, thus indicating the significance of fibrillin-2 in chondrogenesis. In this study, we investigated the transcriptional regulation of fibrillin-2 in chondrogenic differentiation. Although mRNA expression of fibrillin-1, a highly homologous protein to fibrillin-2, remained almost unchanged during chondrogenesis of mouse ATDC5 cells, fibrillin-2 mRNA expression varied. Fibrillin-2 was highly expressed at the early stage and declined progressively during differentiation. The 5′-flanking region of the fibrillin-2 gene contains potential binding sites for E2F, Runx, AP-2, and Sox transcription factors. The promoter activity of fibrillin-2 decreased markedly following deletion and mutagenesis of the E2F binding site between −143 and −136 bp. Overexpression of E2F1 resulted in a marked increase in its promoter activity, whereas expression of other transcription factors including AP-2α and Runx2 had no effect. The increase in promoter activity by E2F1 was completely suppressed by the coexpression of E2F4. E2F2 and E2F3 had positive effects on the promoter activity. Although ATDC5 cells expressed transcripts for the E2F family genes at all stages of differentiation, the expression profiles differed. E2F1 expression remained almost unchanged, whereas E2F4 expression increased markedly at the late stage of differentiation. These results indicated that coordinated expression of the E2F family is critical for the transcriptional regulation of fibrillin-2 during chondrogenesis. J. Cell. Biochem. 106: 580–588, 2009. © 2009 Wiley-Liss, Inc.
