JNCI J Natl Cancer Inst (2015) 107(9): djv180 doi:10.1093/jnci/djv180 First published online June 24, 2015 Editorial
editorial An Atypical Oncogene Within the Atypical E2Fs Affiliation of authors: Cell Division and Cancer group, Molecular Oncology Programme, Spanish National Cancer Research Centre (CNIO) Madrid, Spain (MÁF, MM). Correspondence to: Marcos Malumbres, PhD, Centro Nacional de Investigaciones Oncológicas (CNIO), Melchor Fernández Almagro 3, E-28029 Madrid, Spain (e-mail: [email protected]).
E2F transcription factors are critical components of the transcriptional machinery that modulates the expression of genes required for DNA synthesis and mitosis during the cell cycle. The mammalian E2F factors have been broadly classified into classical activators (E2F1-3) and repressors (E2F4-6). An additional, recently identified branch of this family is composed of two members, E2F7 and E2F8, which are designated as atypical E2Fs because of their structural differences when compared with canonical E2Fs. These atypical E2Fs harbor two DNAbinding domains instead of one, do not require a dimerization partner (DP) to bind to DNA, and regulate transcription independently of retinoblastoma (RB) (1). E2F7 and E2F8 share many transcriptional targets and can form homo- and heterodimers at their target promoters. Atypical E2Fs are thought to play overlapping roles because, whereas E2f7- or E2f8-null mice developed normally, deletion of both genes caused lethality during midgestation in murine embryos as a consequence of massive apoptosis and vascular defects (2). Both E2F7 and E2F8 are able to regulate angiogenesis and lymphangiogenesis in a cell-cycle independent manner by controlling critical regulators such as VEGF, CCBE1, and FLT4 (3,4), although whether these targets are responsible for the vascular defect in E2F7/8 mutant embryos is not clear at present. So far, the major biological function attributed to atypical E2Fs in cell cycle control is the regulation of polyploidization. This function has been revealed by studies in two endocycling mammalian tissues, trophoblast giant cells (TGCs) in the placenta, and hepatocytes in the liver. During placental development, TGCs increase their ploidy enormously through consecutive cycles of endoreplication, in which cells duplicate their genomes without mitosis. TGCs in E2F7/8-null placentas showed reduced ploidy in the presence of aberrant mitotic figures (5), suggesting that these cells were able to enter mitosis in the absence of these atypical E2Fs (Figure 1). Concomitant loss of E2F7 and E2F8 in the liver also reduced polyploidization of hepatocytes, a cell type in which polyploidization is achieved by defective cytokinesis, although these defects did not compromise the functionality or regenerative capacity of the liver in these mutant mice (5,6). Differential gene expression studies in
these tissues showed increased transcription of mitotic genes in the absence of E2F7/8, suggesting that these atypical factors mostly function as transcriptional repressors. Many of those targets were also targets of classical E2F activators, and, indeed, the defects observed in E2F7/8-null cells could be rescued by deleting the canonical E2F3a and E2F1 factors in placenta and liver, respectively (5–7). It was concluded from these initial studies that the main function of atypical E2Fs is to suppress the activity of canonical E2F activators in specialized cell cycles, whereas they are mostly dispensable for conventional mitotic cycles. Recent work suggests that E2F7/8 factors can indeed repress transcription by recruiting the corepressor C-terminal–binding protein 2 (CtBP2) to E2F consensus sites in gene promoters (8). Despite the essential role of E2F7/8 in vascular function or endocycles, very little is known about the relevance of this subfamily in the proliferation of cancer cells. E2F7 was identified as a direct p53 target potently induced during oncogene-induced senescence (9,10). By repressing mitotic genes, E2F7 was proposed to cooperate with RB in limiting oncogenic transformation, although its potential tumor suppressor function has not been further explored (10). In a separate study, low mRNA levels of E2F7 were associated with platinum resistance and reduced survival in ovarian cancer patients (11). Surprisingly, E2F8 behaved in the opposite direction in the same set of patients, with high levels of E2F8 transcripts associated with poor prognosis (11). E2F8 was also found upregulated both at the mRNA and protein levels in hepatocellular carcinoma (HCC) (12), suggesting an unexpected positive function for this transcription factor in the control of tumor cell proliferation. In this issue of the Journal, Park et al. now describe the oncogenic function and therapeutic value of E2F8 in lung adenocarcinoma and squamous cell carcinoma (13). In these tumor cells, E2F8 enhances cell proliferation and its overexpression is associated with poor prognosis, in agreement with the data previously obtained in ovarian cancer. E2F8 depletion by RNA interference or specific morpholinos prevents the proliferation of lung cancer cell lines both in vitro and in vivo. Intriguingly, E2F8 seems to be dispensable for normal lung epithelial cell proliferation, thus suggesting E2F8 as an attractive new therapeutic target for
Received: June 3, 2015; Accepted: June 5, 2015 © The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: [email protected].
1 of 2
Downloaded from http://jnci.oxfordjournals.org/ at University of British Columbia Library on November 16, 2015
Mónica Álvarez-Fernández, Marcos Malumbres
2 of 2 | JNCI J Natl Cancer Inst, 2015, Vol. 107, No. 9
Normal cells
E2F8
E2F7
E2F7
E2F7 E2F1-3
Cancer cells
E2F1-3
HIF1
E2F8
E2F8
E2F7 E2F1-3
E2F8
Promitotic
VEGFA, CCBE1
CCND1, UHRF1
Mitotic cell cycle
Endocycle (polyploidization)
Angiogenesis Lymphangiogenesis
Tumor cell proliferation
Figure 1. Differential function of atypical E2Fs in normal and cancer cells. E2F7/8 transcription factors function as transcriptional repressors of promitotic genes by counteracting the activation mediated by canonical E2F1-3 transcription factors. They also play critical roles in endocycles, likely by repressing mitotic genes. Atypical E2Fs also play cell cycle–independent functions in angiogenesis and lymphangiogenesis, through transcriptional activation of VEGFA or CCBE1, in some cases by forming activation complexes with HIF1. In cancer cells, E2F8, but not E2F7, may function as an oncogene through the transcriptional activation of at least two targets, the E3 ubiquitin ligase UHFR1 and Cyclin D1, the protein product of the CCND1 gene. In liver tumor cells, E2F8 is able counteract the repression of Cyclin D1 mediated by E2F1-3, although to what extent the oncogenic properties of E2F8 depend on competition for E2F1-3 target sites remains unknown.
lung cancer. Transcription factors are not easily druggable in the clinic, but an alternative approach is to target some of their relevant targets. In the current study (13), Park and colleagues identify the E3 ubiquitin ligase UHRF1 as one of the E2F8 targets required for lung cancer cell proliferation. UHRF1, a known oncogene that induces DNA hypomethylation in cancer (14), is not repressed but transcriptionally induced by E2F8 in these tumor cells (Figure 1). Although it is broadly accepted that atypical E2Fs act as transcriptional repressors, there is also evidence of E2F7/8 working as transcriptional activators. For instance, E2F7/8 may play a role in angiogenesis by binding to the Vegfa promoter in a complex with the hypoxia inducible factor (HIF1) and independently of E2F canonical binding sites (3)(Figure 1). Cyclin D1 has also been described as a direct transcriptional target of E2F8 in HCC (12). Thus, the data presented by Park et al. (13) confirm that E2F8 can work both as transcriptional activator and repressor, likely in a target- and cell type–specific manner. What is unique about E2F8 in tumor cell proliferation? To date, no clear therapeutic strategies targeting other E2F targets have been proposed, likely because of their dual roles as activators or repressors of the cell cycle and the possible toxic effect in normal proliferating cells. The antiproliferative potential of E2F8 inhibition in lung cancer cells is unique, as this factor seems to be dispensable for proliferation of untransformed mitotic cells (13). Whether these findings may also apply to other tumor types deserves further investigation. Because E2F8 has no transactivation domain, it remains to be clarified which additional cofactors are required for this activity and whether E2F8 simply competes for E2F1-3 promoter sites or may function independently of this activity in tumor cells. In addition, which are the critical targets downstream of oncogenic E2F8 is not well established, as rescue assays have not been performed with either cyclin D1 or UHRF1. The identification of additional E2F8-relevant targets will likely be critical to understanding its
pro-oncogenic function. Finally, given the crucial role of E2F8 in angiogenesis and lymphangiogenesis (3,4), further studies to evaluate the contribution of E2F8 to metastasis might bring new therapeutic opportunities for the future.
References 1. Lammens T, Li J, Leone G, De Veylder L. Atypical E2Fs: new players in the E2F transcription factor family. Trends Cell Biol. 2009;19:111–118. 2. Li J, Ran C, Li E, et al. Synergistic function of E2F7 and E2F8 is essential for cell survival and embryonic development. Dev Cell. 2008;14:62–75. 3. Weijts BG, Bakker WJ, Cornelissen PW, et al. E2F7 and E2F8 promote angiogenesis through transcriptional activation of VEGFA in cooperation with HIF1. EMBO J. 2012;31:3871–3884. 4. Weijts BG, van Impel A, Schulte-Merker S, de Bruin A. Atypical E2fs Control Lymphangiogenesis through Transcriptional Regulation of Ccbe1 and Flt4. PLoS One 2013;8:e73693. 5. Chen H-Z, Ouseph MM, Li J, et al. Canonical and atypical E2Fs regulate the mammalian endocycle. Nat Cell Biol. 2012;14:1192–1202. 6. Pandit SK, Westendorp B, Nantasanti S, et al. E2F8 is essential for polyploidization in mammalian cells. Nat Cell Biol. 2012;14:1181–1191. 7. Ouseph MM, Li J, Chen H-Z, et al. Atypical E2F repressors and activators coordinate placental development. Dev Cell. 2012;22:849–862. 8. Liu B, Shats I, Angus SP, Gatza ML, Nevins JR. Interaction of E2F7 Transcription Factor with E2F1 and C-terminal-binding Protein (CtBP) Provides a Mechanism for E2F7-dependent Transcription Repression. J Biol Chem. 2013;288:24581–24589. 9. Carvajal LA, Hamard P, Tonnessen C, Manfredi JJ. E2F7, a novel target, is upregulated by p53 and mediates DNA damage-dependent transcriptional repression. Genes Dev. 2012;26:1533–1545. 10. Aksoy O, Chicas A, Zeng T, et al. The atypical E2F family member E2F7 couples the p53 and RB pathways during cellular senescence. Genes Dev. 2012;26:1546–1557. 11. Reimer D, Sadr S, Wiedemair A, et al. Clinical relevance of E2F family members in ovarian cancer--an evaluation in a training set of 77 patients. Clin Cancer Res. 2007;13:144–151. 12. Deng Q, Wang Q, Zong W-Y, et al. E2F8 contributes to human hepatocellular carcinoma via regulating cell proliferation. Cancer Res. 2010;70:782–791. 13. Park S-A, James P, Lee JW, López-Giráldez F, Herbst RS, Seok KJ. E2F8 is a novel therapeutic target for lung cancer. J Natl Cancer Inst. 2015;107(9):djv151 doi:10.1093/jnci/djv151. 14. Mudbhary R, Hoshida Y, Chernyavskaya Y, et al. UHRF1 overexpression drives DNA hypomethylation and hepatocellular carcinoma. Cancer Cell. 2014;25:196–209.
Downloaded from http://jnci.oxfordjournals.org/ at University of British Columbia Library on November 16, 2015
Promitotic
editorial An Atypical Oncogene Within the Atypical E2Fs Affiliation of authors: Cell Division and Cancer group, Molecular Oncology Programme, Spanish National Cancer Research Centre (CNIO) Madrid, Spain (MÁF, MM). Correspondence to: Marcos Malumbres, PhD, Centro Nacional de Investigaciones Oncológicas (CNIO), Melchor Fernández Almagro 3, E-28029 Madrid, Spain (e-mail: [email protected]).
E2F transcription factors are critical components of the transcriptional machinery that modulates the expression of genes required for DNA synthesis and mitosis during the cell cycle. The mammalian E2F factors have been broadly classified into classical activators (E2F1-3) and repressors (E2F4-6). An additional, recently identified branch of this family is composed of two members, E2F7 and E2F8, which are designated as atypical E2Fs because of their structural differences when compared with canonical E2Fs. These atypical E2Fs harbor two DNAbinding domains instead of one, do not require a dimerization partner (DP) to bind to DNA, and regulate transcription independently of retinoblastoma (RB) (1). E2F7 and E2F8 share many transcriptional targets and can form homo- and heterodimers at their target promoters. Atypical E2Fs are thought to play overlapping roles because, whereas E2f7- or E2f8-null mice developed normally, deletion of both genes caused lethality during midgestation in murine embryos as a consequence of massive apoptosis and vascular defects (2). Both E2F7 and E2F8 are able to regulate angiogenesis and lymphangiogenesis in a cell-cycle independent manner by controlling critical regulators such as VEGF, CCBE1, and FLT4 (3,4), although whether these targets are responsible for the vascular defect in E2F7/8 mutant embryos is not clear at present. So far, the major biological function attributed to atypical E2Fs in cell cycle control is the regulation of polyploidization. This function has been revealed by studies in two endocycling mammalian tissues, trophoblast giant cells (TGCs) in the placenta, and hepatocytes in the liver. During placental development, TGCs increase their ploidy enormously through consecutive cycles of endoreplication, in which cells duplicate their genomes without mitosis. TGCs in E2F7/8-null placentas showed reduced ploidy in the presence of aberrant mitotic figures (5), suggesting that these cells were able to enter mitosis in the absence of these atypical E2Fs (Figure 1). Concomitant loss of E2F7 and E2F8 in the liver also reduced polyploidization of hepatocytes, a cell type in which polyploidization is achieved by defective cytokinesis, although these defects did not compromise the functionality or regenerative capacity of the liver in these mutant mice (5,6). Differential gene expression studies in
these tissues showed increased transcription of mitotic genes in the absence of E2F7/8, suggesting that these atypical factors mostly function as transcriptional repressors. Many of those targets were also targets of classical E2F activators, and, indeed, the defects observed in E2F7/8-null cells could be rescued by deleting the canonical E2F3a and E2F1 factors in placenta and liver, respectively (5–7). It was concluded from these initial studies that the main function of atypical E2Fs is to suppress the activity of canonical E2F activators in specialized cell cycles, whereas they are mostly dispensable for conventional mitotic cycles. Recent work suggests that E2F7/8 factors can indeed repress transcription by recruiting the corepressor C-terminal–binding protein 2 (CtBP2) to E2F consensus sites in gene promoters (8). Despite the essential role of E2F7/8 in vascular function or endocycles, very little is known about the relevance of this subfamily in the proliferation of cancer cells. E2F7 was identified as a direct p53 target potently induced during oncogene-induced senescence (9,10). By repressing mitotic genes, E2F7 was proposed to cooperate with RB in limiting oncogenic transformation, although its potential tumor suppressor function has not been further explored (10). In a separate study, low mRNA levels of E2F7 were associated with platinum resistance and reduced survival in ovarian cancer patients (11). Surprisingly, E2F8 behaved in the opposite direction in the same set of patients, with high levels of E2F8 transcripts associated with poor prognosis (11). E2F8 was also found upregulated both at the mRNA and protein levels in hepatocellular carcinoma (HCC) (12), suggesting an unexpected positive function for this transcription factor in the control of tumor cell proliferation. In this issue of the Journal, Park et al. now describe the oncogenic function and therapeutic value of E2F8 in lung adenocarcinoma and squamous cell carcinoma (13). In these tumor cells, E2F8 enhances cell proliferation and its overexpression is associated with poor prognosis, in agreement with the data previously obtained in ovarian cancer. E2F8 depletion by RNA interference or specific morpholinos prevents the proliferation of lung cancer cell lines both in vitro and in vivo. Intriguingly, E2F8 seems to be dispensable for normal lung epithelial cell proliferation, thus suggesting E2F8 as an attractive new therapeutic target for
Received: June 3, 2015; Accepted: June 5, 2015 © The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: [email protected].
1 of 2
Downloaded from http://jnci.oxfordjournals.org/ at University of British Columbia Library on November 16, 2015
Mónica Álvarez-Fernández, Marcos Malumbres
2 of 2 | JNCI J Natl Cancer Inst, 2015, Vol. 107, No. 9
Normal cells
E2F8
E2F7
E2F7
E2F7 E2F1-3
Cancer cells
E2F1-3
HIF1
E2F8
E2F8
E2F7 E2F1-3
E2F8
Promitotic
VEGFA, CCBE1
CCND1, UHRF1
Mitotic cell cycle
Endocycle (polyploidization)
Angiogenesis Lymphangiogenesis
Tumor cell proliferation
Figure 1. Differential function of atypical E2Fs in normal and cancer cells. E2F7/8 transcription factors function as transcriptional repressors of promitotic genes by counteracting the activation mediated by canonical E2F1-3 transcription factors. They also play critical roles in endocycles, likely by repressing mitotic genes. Atypical E2Fs also play cell cycle–independent functions in angiogenesis and lymphangiogenesis, through transcriptional activation of VEGFA or CCBE1, in some cases by forming activation complexes with HIF1. In cancer cells, E2F8, but not E2F7, may function as an oncogene through the transcriptional activation of at least two targets, the E3 ubiquitin ligase UHFR1 and Cyclin D1, the protein product of the CCND1 gene. In liver tumor cells, E2F8 is able counteract the repression of Cyclin D1 mediated by E2F1-3, although to what extent the oncogenic properties of E2F8 depend on competition for E2F1-3 target sites remains unknown.
lung cancer. Transcription factors are not easily druggable in the clinic, but an alternative approach is to target some of their relevant targets. In the current study (13), Park and colleagues identify the E3 ubiquitin ligase UHRF1 as one of the E2F8 targets required for lung cancer cell proliferation. UHRF1, a known oncogene that induces DNA hypomethylation in cancer (14), is not repressed but transcriptionally induced by E2F8 in these tumor cells (Figure 1). Although it is broadly accepted that atypical E2Fs act as transcriptional repressors, there is also evidence of E2F7/8 working as transcriptional activators. For instance, E2F7/8 may play a role in angiogenesis by binding to the Vegfa promoter in a complex with the hypoxia inducible factor (HIF1) and independently of E2F canonical binding sites (3)(Figure 1). Cyclin D1 has also been described as a direct transcriptional target of E2F8 in HCC (12). Thus, the data presented by Park et al. (13) confirm that E2F8 can work both as transcriptional activator and repressor, likely in a target- and cell type–specific manner. What is unique about E2F8 in tumor cell proliferation? To date, no clear therapeutic strategies targeting other E2F targets have been proposed, likely because of their dual roles as activators or repressors of the cell cycle and the possible toxic effect in normal proliferating cells. The antiproliferative potential of E2F8 inhibition in lung cancer cells is unique, as this factor seems to be dispensable for proliferation of untransformed mitotic cells (13). Whether these findings may also apply to other tumor types deserves further investigation. Because E2F8 has no transactivation domain, it remains to be clarified which additional cofactors are required for this activity and whether E2F8 simply competes for E2F1-3 promoter sites or may function independently of this activity in tumor cells. In addition, which are the critical targets downstream of oncogenic E2F8 is not well established, as rescue assays have not been performed with either cyclin D1 or UHRF1. The identification of additional E2F8-relevant targets will likely be critical to understanding its
pro-oncogenic function. Finally, given the crucial role of E2F8 in angiogenesis and lymphangiogenesis (3,4), further studies to evaluate the contribution of E2F8 to metastasis might bring new therapeutic opportunities for the future.
References 1. Lammens T, Li J, Leone G, De Veylder L. Atypical E2Fs: new players in the E2F transcription factor family. Trends Cell Biol. 2009;19:111–118. 2. Li J, Ran C, Li E, et al. Synergistic function of E2F7 and E2F8 is essential for cell survival and embryonic development. Dev Cell. 2008;14:62–75. 3. Weijts BG, Bakker WJ, Cornelissen PW, et al. E2F7 and E2F8 promote angiogenesis through transcriptional activation of VEGFA in cooperation with HIF1. EMBO J. 2012;31:3871–3884. 4. Weijts BG, van Impel A, Schulte-Merker S, de Bruin A. Atypical E2fs Control Lymphangiogenesis through Transcriptional Regulation of Ccbe1 and Flt4. PLoS One 2013;8:e73693. 5. Chen H-Z, Ouseph MM, Li J, et al. Canonical and atypical E2Fs regulate the mammalian endocycle. Nat Cell Biol. 2012;14:1192–1202. 6. Pandit SK, Westendorp B, Nantasanti S, et al. E2F8 is essential for polyploidization in mammalian cells. Nat Cell Biol. 2012;14:1181–1191. 7. Ouseph MM, Li J, Chen H-Z, et al. Atypical E2F repressors and activators coordinate placental development. Dev Cell. 2012;22:849–862. 8. Liu B, Shats I, Angus SP, Gatza ML, Nevins JR. Interaction of E2F7 Transcription Factor with E2F1 and C-terminal-binding Protein (CtBP) Provides a Mechanism for E2F7-dependent Transcription Repression. J Biol Chem. 2013;288:24581–24589. 9. Carvajal LA, Hamard P, Tonnessen C, Manfredi JJ. E2F7, a novel target, is upregulated by p53 and mediates DNA damage-dependent transcriptional repression. Genes Dev. 2012;26:1533–1545. 10. Aksoy O, Chicas A, Zeng T, et al. The atypical E2F family member E2F7 couples the p53 and RB pathways during cellular senescence. Genes Dev. 2012;26:1546–1557. 11. Reimer D, Sadr S, Wiedemair A, et al. Clinical relevance of E2F family members in ovarian cancer--an evaluation in a training set of 77 patients. Clin Cancer Res. 2007;13:144–151. 12. Deng Q, Wang Q, Zong W-Y, et al. E2F8 contributes to human hepatocellular carcinoma via regulating cell proliferation. Cancer Res. 2010;70:782–791. 13. Park S-A, James P, Lee JW, López-Giráldez F, Herbst RS, Seok KJ. E2F8 is a novel therapeutic target for lung cancer. J Natl Cancer Inst. 2015;107(9):djv151 doi:10.1093/jnci/djv151. 14. Mudbhary R, Hoshida Y, Chernyavskaya Y, et al. UHRF1 overexpression drives DNA hypomethylation and hepatocellular carcinoma. Cancer Cell. 2014;25:196–209.
Downloaded from http://jnci.oxfordjournals.org/ at University of British Columbia Library on November 16, 2015
Promitotic
